Novel Inhibitors of Glutaminyl Cyclase

ABSTRACT

The present invention relates to novel inhibitors of glutaminyl cyclase and combinations thereof for the treatment of neuronal disorders, especially Alzheimer&#39;s disease, Down Syndrome, Parkinson disease, Chorea Huntington, pathogenic psychotic conditions, schizophrenia, impaired food intake, sleep-wakefulness, impaired homeostatic regulation of energy metabolism, impaired autonomic function, impaired hormonal balance, impaired regulation, body fluids, hypertension, fever, sleep dysregulation, anorexia, anxiety related disorders including depression, seizures including epilepsy, drug withdrawal and alcoholism, neurodegenerative disorders including cognitive dysfunction and dementia.

RELATED APPLICATIONS

This application is a Continuation application of U.S. patentapplication Ser. No. 11/051,760, filed Feb. 4, 2005, which in turn was aContinuation-in-Part application of U.S. patent application Ser. No.10/838,993 filed May 5, 2004, which in turn claims the benefit of U.S.Provisional Application Ser. No. 60/542,133, filed Feb. 5, 2004, andU.S. Provisional Application Ser. No. 60/634,364, filed Dec. 8, 2004.Each of the above applications are incorporated herein by reference intheir entirety.

FIELD OF THE INVENTION

The invention relates to glutaminyl cyclase (QC, EC 2.3.2.5) thatcatalyzes the intramolecular cyclization of N-terminal glutamineresidues into pyroglutamic acid (5-oxo-prolyl, pGlu*) under liberationof ammonia and the intramolecular cyclization of N-terminal glutamateresidues into pyroglutamic acid under liberation of water.

BACKGROUND OF THE INVENTION

Glutaminyl cyclase (QC, EC 2.3.2.5) catalyzes the intramolecularcyclization of N-terminal glutamine residues into pyroglutamic acid(pGlu*) liberating ammonia. A QC was first isolated by Messer from thelatex of the tropical plant Carica papaya in 1963 (Messer, M. 1963Nature 4874, 1299). 24 years later, a corresponding enzymatic activitywas discovered in animal pituitary (Busby, W. H. J. et al. 1987 J BiolChem 262, 8532-8536; Fischer, W. H. and Spiess, J. 1987 Proc Natl AcadSci USA 84, 3628-3632). For the mammalian QC, the conversion of Gln intopGlu by QC could be shown for the precursors of TRH and GnRH (Busby, W.H. J. et al. 1987 J Biol Chem 262, 8532-8536; Fischer, W. H. and Spiess,J. 1987 Proc Natl Acad Sci USA 84, 3628-3632). In addition, initiallocalization experiments of QC revealed a co-localization with itsputative products of catalysis in bovine pituitary, further improvingthe suggested function in peptide hormone synthesis (Bockers, T. M. etal. 1995 J Neuroendocrinol 7, 445-453). In contrast, the physiologicalfunction of the plant QC is less clear. In the case of the enzyme fromC. papaya, a role in the plant defense against pathogenic microorganismswas suggested (E I Moussaoui, A. et al. 2001 Cell Mol Life Sci 58,556-570). Putative QCs from other plants were identified by sequencecomparisons recently (Dahl, S. W. et al. 2000 Protein Expr Purif 20,27-36). The physiological function of these enzymes, however, is stillambiguous.

The QCs known from plants and animals show a strict specificity forL-Glutamine in the N-terminal position of the substrates and theirkinetic behavior was found to obey the Michaelis-Menten equation (Pohl,T. et al. 1991 Proc Natl Acad Sci USA 88, 10059-10063; Consalvo, A. P.et al. 1988 Anal Biochem 175, 131-138; Gololobov, M. Y. et al. 1996 BiolChem Hoppe Seyler 377, 395-398). A comparison of the primary structuresof the QCs from C. papaya and that of the highly conserved QC frommammals, however, did not reveal any sequence homology (Dahl, S. W. etal. 2000 Protein Expr Purif 20, 27-36). Whereas the plant QCs appear tobelong to a new enzyme family (Dahl, S. W. et al. 2000 Protein ExprPurif 20, 27-36), the mammalian QCs were found to have a pronouncedsequence homology to bacterial aminopeptidases (Bateman, R. C. et al.2001 Biochemistry 40, 11246-11250), leading to the conclusion that theQCs from plants and animals have different evolutionary origins.

Recently, it was shown that recombinant human QC as well as QC-activityfrom brain extracts catalyze both, the N-terminal glutaminyl as well asglutamate cyclization. Most striking is the finding, thatcyclase-catalyzed Glu₁-conversion is favored around pH 6.0 whileGln₁-conversion to pGlu-derivatives occurs with a pH-optimum of around8.0. Since the formation of pGlu-Aβ-related peptides can be suppressedby inhibition of recombinant human QC and QC-activity from pig pituitaryextracts, the enzyme QC is a target in drug development for treatment ofAlzheimer's disease.

EP 02 011 349.4 discloses polynucleotides encoding insect glutaminylcyclase, as well as polypeptides encoded thereby. This applicationfurther provides host cells comprising expression vectors comprisingpolynucleotides of the invention. Isolated polypeptides and host cellscomprising insect QC are useful in methods of screening for agents thatreduce glutaminyl cyclase activity. Such agents are useful aspesticides.

DEFINITIONS Enzyme Inhibitors

Reversible enzyme inhibitors: comprise competitive inhibitors,non-competitive reversible inhibitors, slow-binding or tight-bindinginhibitors, transition state analogs and multisubstrate analogs.

Competitive inhibitors show

-   -   i) non-covalent interactions with the enzyme,    -   ii) compete with substrate for the enzyme active site,

The principal mechanism of action of a reversible enzyme inhibitor andthe definition of the dissociation constant can be visualized asfollows:

The formation of the enzyme-inhibitor [E−I] complex prevents binding ofsubstrates, therefore the reaction cannot proceed to the normalphysiological product, P. A larger inhibitor concentration [I] leads tolarger [E−I], leaving less free enzyme to which the substrate can bind.

Non-Competitive Reversible Inhibitors

-   -   i) bind at a site other than active site (allosteric binding        site)    -   ii) cause a conformational change in the enzyme which decreases        or stops catalytic activity.

Slow-Binding or Tight-Binding Inhibitors

-   -   i) are competitive inhibitors where the equilibrium between        inhibitor and enzyme is reached slowly,    -   ii) (k_(on) is slow), possibly due to conformational changes        that must occur in the enzyme or inhibitor        -   a) are often transition state analogs        -   b) are effective at concentrations similar to the enzyme            conc. (subnanomolar K_(D) values)        -   c) due to k_(off) values being so low these types of            inhibitors are “almost” irreversible

Transition State Analogs

are competitive inhibitors which mimic the transition state of an enzymecatalyzed reaction. Enzyme catalysis occurs due to a lowering of theenergy of the transition state, therefore, transition state binding isfavored over substrate binding.

Multisubstrate Analogs

For a reaction involving two or more substrates, a competitive inhibitoror transition state analog can be designed which contains structuralcharacteristics resembling two or more of the substrates.

Irreversible enzyme inhibitors: drive the equilibrium between theunbound enzyme and inhibitor and enzyme inhibitor complex (E+I<--->E−I)all the way to the right with a covalent bond (˜100 kcal/mole), makingthe inhibition irreversible.

Affinity Labeling Agents

-   -   Active-site directed irreversible inhibitors (competitive        irreversible inhibitor) are recognized by the enzyme        (reversible, specific binding) followed by covalent bond        formation, and        -   i) are structurally similar to substrate, transition state            or product allowing for specific interaction between drug            and target enzyme,        -   ii) contain reactive functional group (e.g. a nucleophile,            —COCH₂Br) allowing for covalent bond formation        -   The reaction scheme below describes an active-site directed            reagent with its target enzyme where K_(D) is the            dissociation constant and k_(inactivation) is the rate of            covalent bond formation.

-   -   Mechanism-based enzyme inactivators (also called suicide        inhibitors) are active-site directed reagents (unreactive) which        binds to the enzyme active site where it is transformed to a        reactive form (activated) by the enzyme's catalytic        capabilities. Once activated, a covalent bond between the        inhibitor and the enzyme is formed.    -   The reaction scheme below shows the mechanism of action of a        mechanism based enzyme inactivator, where K_(D) is the        dissociation complex, k₂ is the rate of activation of the        inhibitor once bound to the enzyme, k₃ is the rate of        dissociation of the activated inhibitor, P, from the enzyme        (product can still be reactive) from the enzyme and k₄ is the        rate of covalent bond formation between the activated inhibitor        and the enzyme.

-   -   Inactivation (covalent bond formation, k₄) must occur prior to        dissociation (k₃) otherwise the now reactive inhibitor is        released into the environment. Partition ratio, k₃/k₄: ratio of        released product to inactivation should be minimized for        efficient inactivation of the system and minimal undesirable        side reactions. A large partition ratio (favors dissocation)        leads to nonspecific reactions.

Uncompetitive enzyme inhibitors: From the definition of uncompetitiveinhibitor (an inhibitor which binds only to ES complexes) the followingequilibria can be written:

The ES complex dissociates the substrate with a dissociation constantequal to Ks, whereas the ESI complex does not dissociate it (i.e has aKs value equal to zero). The K_(m)'s of Michaelis-Menten type enzymesare expected to be reduced. Increasing substrate concentration leads toincreasing ESI concentration (a complex incapable of progressing toreaction products), therefore the inhibit ion can not be removed.

Preferred according to the present invention are competitive enzymeinhibitors. Most preferred are competitive reversible enzyme inhibitors.

The terms “k_(i)” or “K_(i)” and “K_(D)” are binding constants, whichdescribe the binding of an inhibitor to and the subsequent release froman enzyme. Another measure is the “IC₅₀” value, which reflects theinhibitor concentration which at a given substrate concentration resultsin 50% enzyme activity.

The term “DP IV-inhibitor” or “dipeptidyl peptidase IV inhibitor” isgenerally known to a person skilled in the art and means enzymeinhibitors, which inhibit the catalytic activity of DP IV or DP IV-likeenzymes.

“DP IV-activity” is defined as the catalytic activity of dipeptidylpeptidase IV (DP IV) and DP IV-like enzymes. These enzymes arepost-proline (to a lesser extent post-alanine, post-serine orpost-glycine) cleaving serine proteases found in various tissues of thebody of a mammal including kidney, liver, and intestine, where theyremove dipeptides from the N-terminus of biologically active peptideswith a high specificity when proline or alanine form the residues thatare adjacent to the N-terminal amino acid in their sequence.

The term “PEP-inhibitor” or “prolyl endopeptidase inhibitor” isgenerally known to a person skilled in the art and means enzymeinhibitors, which inhibit the catalytic activity of prolyl endopeptidase(PEP, prolyl oligopeptidase, POP).

“PEP-activity” is defined as the catalytic activity of an endoproteasethat is capable to hydrolyze post proline bonds in peptides or proteinswere the proline is in amino acid position 3 or higher counted from theN-terminus of a peptide or protein substrate.

The term “QC” as used herein comprises glutaminyl cyclase (QC) andQC-like enzymes. QC and QC-like enzymes have identical or similarenzymatic activity, further defined as QC activity. In this regard,QC-like enzymes can fundamentally differ in their molecular structurefrom QC.

The term “QC activity” as used herein is defined as intramolecularcyclization of N-terminal glutamine residues into pyroglutamic acid(pGlu*) or of N-terminal L-homoglutamine or L-β-homoglutamine to acyclic pyro-homoglutamine derivative under liberation of ammonia. Seetherefore schemes 1 and 2.

The term “EC” as used herein comprises the side activity of QC andQC-like enzymes as glutamate cyclase (EC), further defined as ECactivity.

The term “EC activity” as used herein is defined as intramolecularcyclization of N-terminal glutamate residues into pyroglutamic acid(pGlu*) by QC. See therefore scheme 3.

The term “QC-inhibitor” “glutaminyl cyclase inhibitor” is generallyknown to a person skilled in the art and means enzyme inhibitors, whichinhibit the catalytic activity of glutaminyl cyclase (QC) or itsglutamyl cyclase (EC) activity.

Potency of QC Inhibition

In light of the correlation with QC inhibition, in preferredembodiments, the subject method and medical use utilize an agent with aKi for QC inhibition of 10 μM or less, more preferably of 1 μM or less,even more preferably of 0.1 μM or less or 0.01 μM or less, or mostpreferably 0.01 μM or less. Indeed, inhibitors with Ki values in thelower micromolar, preferably the nanomolar and even more preferably thepicomolar range are contemplated. Thus, while the active agents aredescribed herein, for convenience, as “QC inhibitors”, it will beunderstood that such nomenclature is not intending to limit the subjectof the invention to a particular mechanism of action.

Molecular Weight of QC Inhibitors

In general, the QC inhibitors of the subject method or medical use willbe small molecules, e.g., with molecular weights of 1000 g/mole or less,500 g/mole or less, preferably of 400 g/mole or less, and even morepreferably of 350 g/mole or less and even of 300 g/mole or less.

The term “subject” as used herein, refers to an animal, preferably amammal, most preferably a human, who has been the object of treatment,observation or experiment.

The term “therapeutically effective amount” as used herein, means thatamount of active compound or pharmaceutical agent that elicits thebiological or medicinal response in a tissue system, animal or humanbeing sought by a researcher, veterinarian, medical doctor or otherclinician, which includes alleviation of the symptoms of the disease ordisorder being treated.

As used herein, the term “pharmaceutically acceptable” embraces bothhuman and veterinary use: for example the term “pharmaceuticallyacceptable” embraces a veterinarily acceptable compound or a compoundacceptable in human medicine and health care.

Throughout the description and the claims the expression “acyl”, unlessspecifically limited, denotes a C₁₋₁₂ acyl residue, preferably a C₁₋₈acyl residue and especially preferred a C₁₋₄ acyl residue. Examples ofacyl include alkanoyl groups mentioned below and benzoyl.

“Peptides” are selected from dipeptides to decapeptides, preferred aredipeptides, tripeptides, tetrapeptides and pentapeptides. The aminoacids for the formation of the “peptides” can be selected from thoselisted below.

Throughout the description and the claims the expression “alkyl”, unlessspecifically limited, denotes a C₁₋₁₂ alkyl group, preferably a C₁₋₆alkyl group. Alkyl groups may be straight chain or branched. Suitablealkyl groups include, for example, methyl, ethyl, propyl (e.g. n-propyland isopropyl), (n-butyl, tert-butyl and sec-butyl), pentyl, hexyl,heptyl (e.g. n-heptyl) and octyl (e.g. n-octyl). The expression “alk”,for example in the expression “alkoxy”, and the expression “alkan”, forexample in the expression “alkanoyl”, should be interpreted inaccordance with the definition of “alkyl”. Exemplary alkoxy groupsinclude methoxy, ethoxy, butoxy (e.g. n-butoxy), heptyloxy (e.g.n-heptyloxy) and octyloxy (e.g. n-octyloxy). Exemplary alkanoyl (i.e.acyl groups) include ethanoyl (i.e. acetyl), propionyl and butyryl.

The expression “alkenyl”, unless specifically limited, denotes a C₂₋₁₂alkenyl group, preferably a C₂₋₆ alkenyl group, which contains at leastone double bond at any desired location. Alkenyl groups may be straightchain or branched. Exemplary alkenyl groups include ethenyl, propenyland butenyl.

The expression “alkynyl”, unless specifically limited, denotes a C₂₋₁₂alkynyl group, preferably a C₂₋₆ alkynyl group, which contains at leastone triple bond at any desired location. Alkynyl groups may be straightchain or branched. Exemplary alkenyl groups include ethynyl, propynyland butynyl.

The expression “cycloalkyl”, unless specifically limited, denotes aC₃₋₁₂ cycloalkyl group, preferably a C₃₋₈ cycloalkyl group. Exemplarycycloalkyl groups include cylcopropyl, cyclobutyl, cyclopropyl,cyclohexyl, cycloheptyl and cyclooctyl. Cycloalkyl groups may bebranched in which case the number of carbons indicates the total numberof carbons in the moiety.

The expression “heterocyclic”, unless specifically limited, denotes acycloalkyl residue, wherein one or more (e.g. 1, 2 or 3) ring atoms arereplaced by heteroatoms selected from N, S or O. Exemplary heterocyclicgroups containing one hetero atom include pyrrolidine, tetrahydrofuranand piperidine. Such groups may be optionally substituted eg by alkyl,oxo or hydroxyl.

Concrete examples of a heterocyclic group comprise a substituted orunsubstituted oxirano, aziridino, oxacyclopropyl, azacyclopropyl,thiirano, oxetano, thietano, pyrrolidino, tetrahydrofurano, thiolano,1,1-dioxo-thiolano, 1,3-dioxolano, thiazolidino, imidazolidino,oxazolidino, pyrazolidino, tetrahydropyrano, piperidino, urotropino,piperazino, N-methyl-piperazino, (2-(N-methyl)-N′-piperazinyl)-ethyl,(4N-(2′-hydroxyethyl)-1N-piperazinyl),(2-(4N-(2′-hydroxyethyl)-1N-piperazinyl)-ethyloxy), morpholino,2-(N-morpholino)-ethyl group, as well as lactams, lactones, cyclicimides and cyclic anhydrides.

The expression “carbocylic”, unless specifically limited, denotes acarbocylic group containing between 3 and 12 carbon atoms, morepreferably between 3 and 8 carbon atoms. A carbocylic group, as usedherein, refers to a group other than aryl or cycloalkyl which comprisesat least one ring of carbon atoms without heteroatoms. Examples ofcarbocylic groups include bridged ring systems (e.g.bicyclo[2.2.1]heptenyl) and partially unsaturated ring systems.

The expression “aryl”, unless specifically limited, denotes a C₆₋₁₂ arylgroup, preferably a C₆₋₈ aryl group. Aryl groups will contain at leastone aromatic ring (e.g. one, two or three rings), but may also comprisepartially or fully unsaturated rings. An example of an aryl group withone aromatic ring is phenyl. Examples of aromatic groups with twoaromatic rings include naphyl. Examples of aryl groups which containpartially or fully unsaturated rings include pentalene and indene. Asnoted below, aryl groups may optionally be substituted. Further examplesfor aryl groups are 4-fluoro-phenyl, 3-fluoro-phenyl,pentafluoro-phenyl, 4-hydroxyphenyl-, 3-nitro-phenyl-,4-(trifluoromethyl)-phenyl-, 4-anilinyl-, 2-biphenylyl-, 3-biphenylyl-,4-biphenylyl-, indenyl-, 1-naphthyl-, or 2-naphthyl-, 1-anthracenyl-,2-anthracenyl-, 3-anthracenyl-groups.

Examples of -alkylaryl include phenylmethyl-(benzyl) and phenylethyl,2-phenyleth-1-yl, p-tolyl-methyl-, p-tolyl-ethyl-, m-tolyl-methyl-,m-tolyl-ethyl-, o-tolyl-methyl-, o-tolyl-ethyl-,2-(4-ethyl-phenyl)-eth-1-yl-, 2,3-dimethyl-phenyl-methyl-,2,4-dimethyl-phenyl-methyl-, 2,5-dimethyl-phenyl-methyl-,2,6-dimethyl-phenyl-methyl-, 3,4-dimethyl-phenyl-methyl-,3,5-dimethyl-phenyl-methyl-, 2,4,6-trimethyl-phenyl-methyl-,2,3-dimethyl-phenyl-ethyl-, 2,4-dimethyl-phenyl-ethyl-,2,5-dimethyl-phenyl-ethyl-, 2,6-dimethyl-phenyl-ethyl-,3,4-dimethyl-phenyl-ethyl-, 3,5-dimethyl-phenyl-ethyl-,2,4,6-trimethyl-phenyl-ethyl-, benzhydryl (=diphenyl-methyl), benzhydryl(=diphenyl-ethyl), trityl (=triphenyl-methyl), trityl(=triphenyl-ethyl), α-styryl, α-styryl, cumyl, 2-ethyl-phenyl-methyl-,3-ethyl-phenyl-methyl-, 4-ethyl-phenyl-methyl-, 2-ethyl-phenyl-ethyl-,3-ethyl-phenyl-ethyl-, 4-ethyl-phenyl-ethyl-, 2-fluoro-benzyl,1-methyl-2-fluoro-phen-6-yl-methyl-,1-methyl-2-fluoro-phen-4-yl-methyl-, 1-methyl-2-fluoro-phen-6-yl-ethyl-,1-methyl-2-fluoro-phen-4-yl-ethyl-, 1H-indenyl-methyl-,2H-indenyl-methyl-, 1H-indenyl-ethyl-, 2H-indenyl-ethyl-,indanyl-methyl-, indan-1-on-2-yl-methyl-, indan-1-on-2-yl-ethyl-,tetralinyl-methyl-, tetralinyl-ethyl-, fluorenyl-methyl-,fluorenyl-ethyl-, (3-phenyl)-cyclopent-1-yl ?,dihydronaphthalinyl-methyl-, dihydronaphthalinyl-ethyl-, or(4-cyclohexyl)-phenyl-methyl-, (4-cyclohexyl)-phenyl-ethyl-.

The expression “heteroaryl”, unless specifically limited, denotes as anaryl residue, wherein one or more (e.g. 1, 2, 3, or 4, preferably 1, 2or 3) ring atoms are replaced by heteroatoms selected from N, S and O orelse a 5-membered aromatic ring containing one or more (e.g. 1, 2, 3, or4, preferably 1, 2 or 3) ring atoms selected from N, S and O. As notedbelow, heteroaryl groups may optionally be substituted. Exemplaryheteroaryl groups include, pyridine (eg 2, 3 or 4-pyridine), pyrimidine,quinoline, pyrrole, furan, thiophene, oxazole, pyrazole, benzodioxolane,benzodioxane, benzothiophene, benzodioxepine, and thiazolyl,1-imidazolyl, 2-imidazolyl, 4-imidazolyl, 3-phenyl-1-pyrrolyl,isoxazolyl, isothiazolyl, 3-pyrazolyl, 1,2,3-triazolyl, 1,2,4-triazolyl,tetrazolyl, pyridazinyl, pyrazinyl, indazolyl, 6-indolyl,benzimidazolyl, isochinolinyl, purinyl, carbazolinyl, acridinyl, and2,3′-bifuryl groups. Examples of -alkylheteroaryl includepyridinylmethyl-, N-methyl-pyrrol-2-methyl-N-methyl-pyrrol-2-ethyl-,N-methyl-pyrrol-3-methyl-, N-methyl-pyrrol-3-ethyl-,2-methyl-pyrrol-1-methyl-, 2-methyl-pyrrol-1-ethyl-,3-methyl-pyrrol-1-methyl-, 3-methyl-pyrrol-1-ethyl-, 4-pyridino-methyl-,4-pyridino-ethyl-, 2-(thiazol-2-yl)-ethyl-,tetrahydroisochinolinyl-methyl-, tetrahydroisochinolinyl-ethyl-,2-ethyl-indol-1-methyl-, 2-ethyl-indol-1-ethyl-,3-ethyl-indol-1-methyl-, 3-ethyl-indol-1-ethyl-,4-methyl-pyridin-2-methyl-, 4-methyl-pyridin-2-yl-ethyl-,4-methyl-pyridin-3-methyl, 4-methyl-pyridin-3-ethyl.

The aforementioned aryl and heteroaryl groups may, where appropriate,optionally be substituted.

The expression “substitution” or “substituted” includes the substitutionby one or more (e.g. 1, 2 or 3, preferably 1 or 2) monovalent ormultivalent functional groups. Suitable substituent groups includealkyl, cycloalkyl, aryl (eg phenyl), heteroaryl (eg furyl), carbocylic,heterocyclic, alkoxy, cycloalkoxy, aryloxy, heteroaryloxy,carbocyclicoxy, heterocyclicoxy, alkenyloxy, alkynyloxy, alkenyl,alkynyl, acyl, alkanoyl, alkoxyalkanoyl, alkoxyalkyl, heteroarylalkyl,arylalkyl, arylalkyloxy, heteroarylalkyloxy, nitro, —S-alkyl (e.g.methylthio) halo (e.g. fluoro, chloro, bromo and iodo), cyano, hydroxyl,—SO₂alkyl, —SO₂aryl, —SO₂heteroaryl, —SO₂cycloalkyl —SO₂heterocyclic,—CO₂H, —CO₂alkyl, —NH₂, —NHalkyl, —N(alkyl)₂ (e.g. dimethylamino),—CO—N(alkyl)₂ and —CO—NH(alkyl).

Alkyl groups including derivatives such as alkoxy together with alkenyl,alkynyl and cycloalkyl groups may optionally be halogen substituted e.g.substituted by fluoro. For example, halo substituted alkyl groupsinclude trifluoromethyl and halo substituted alkoxy groups includetrifluoromethoxy.

The term “halogen” comprises fluorine (—F), chlorine (—Cl), bromine(—Br), and iodine (—I), respectively.

Amino acids which can be used in the present invention are L and D-aminoacids, N-alkylated amino acids, N-methyl-amino acids, aza-amino acids;allo- and threo-forms of Ile and Thr, which can, e.g. be α-, β- orω-amino acids, whereof α-amino acids are preferred.

Examples of amino acids are:

aspartic acid (Asp), glutamic acid (Glu), arginine (Arg), lysine (Lys),histidine (His), glycine (Gly), serine (Ser), cysteine (Cys), threonine(Thr), asparagine (Asn), glutamine (Gln), tyrosine (Tyr), alanine (Ala),proline (Pro), valine (Val), isoleucine (Ile), leucine (Leu), methionine(Met), phenylalanine (Phe), tryptophan (Trp), hydroxyproline (Hyp),beta-alanine (beta-Ala), 2-aminooctanoic acid (Aoa),acetidine-(2)-carboxylic acid (Ace), pipecolic acid (Pip),3-aminopropionic acid, 4-aminobutyric acid and so forth,alpha-aminoisobutyric acid (Aib), sarcosine (Sar), ornithine (Orn),citrulline (Cit), homoarginine (Har), t-butylalanine (t-butyl-Ala),t-butylglycine (t-butyl-Gly), N-methylisoleucine (N-Melle),phenylglycine (Phg), cyclohexylalanine (Cha), norleucine (Nle), cysteicacid (Cya) and methionine sulfoxide (MSO), acetyl-Lys, modified aminoacids such as phosphoryl-serine (Ser(P)), benzyl-serine (Ser(Bzl)) andphosphoryl-tyrosine (Tyr(P)), 2-aminobutyric acid (Abu),aminoethylcysteine (AECys), carboxymethylcysteine (Cmc), dehydroalanine(Dha), dehydroamino-2-butyric acid (Dhb), carboxyglutaminic acid (Gla),homoserine (Hse), hydroxylysine (Hyl), cis-hydroxyproline (cis Hyp),trans-hydroxyproline (transHyp), isovaline (Iva), pyroglutamic acid(Pyr), norvaline (Nva), 2-aminobenzoic acid (2-Abz), 3-aminobenzoic acid(3-Abz), 4-aminobenzoic acid (4-Abz), 4-(aminomethyl)benzoic acid (Amb),4-(aminomethyl)cyclohexanecarboxylic acid (4-Amc), Penicillamine (Pen),2-amino-4-cyanobutyric acid (Cba), cycloalkane-carboxylic acids.Examples of 1-amino acids are e.g.: 5-Ara (a minoraleric acid), 6-Ahx(aminohexanoic acid), 8-Aoc (aminooctanoic acid), 9-Anc (aminovanoicacid), 10-Adc (aminodecanoic acid), 11-Aun (aminoundecanoic acid),12-Ado (aminododecanoic acid). Further amino acids are: indanylglycine(Igl), indoline-2-carboxylic acid (ldc), octahydroindole-2-carboxylicacid (Oic), diaminopropionic acid (Dpr), diaminobutyric acid (Dbu),naphtylalanine (1-NaI) and (2-NaI), 4-aminophenylalanine (Phe(4-NH₂)),4-benzoylphenylalanine (Bpa), diphenylalanine (Dip),4-bromophenylalanine (Phe(4-Br)), 2-chlorophenylalanine (Phe(2-Cl)),3-chlorophenylalanine (Phe(3-Cl)), 4-chlorophenylalanine (Phe(4-Cl)),3,4-chlorophenylalanine (Phe (3,4-C₁₂)), 3-fluorophenylalanine(Phe(3-F)), 4-fluorophenylalanine (Phe(4-F)), 3,4-fluorophenylalanine(Phe(3,4-F₂)), pentafluorophenylalanine (Phe(F₅)),4-guanidinophenylalanine (Phe(4-guanidino)), homophenylalanine (hPhe),3-jodophenylalanine (Phe(3-J)), 4-jodophenylalanine (Phe(4-J)),4-methylphenylalanine (Phe(4-Me)), 4-nitrophenylalanine (Phe-4-NO₂)),biphenylalanine (Bip), 4-phosphonomethylphenylalanine (Pmp),cyclohexylglycine (Ghg), 3-pyridinylalanine (3-Pal), 4-pyridinylalanine(4-Pal), 3,4-dehydroproline (A-Pro), 4-ketoproline (Pro(4-keto)),thioproline (Thz), isonipecotic acid (Inp),1,2,3,4,-tetrahydroisoquinolin-3-carboxylic acid (Tic), propargylglycine(Pra), 6-hydroxynorleucine (NU(6-OH)), homotyrosine (hTyr),3-jodotyrosine (Tyr(3-J)), 3,5-dijodotyrosine (Tyr(3,5-J₂)),methyltyrosine (Tyr(Me)), 2′,6′-dimethyltyrosine (Dmt), 3-NO₂-tyrosine(Tyr(3-NO₂)), phosphotyrosine (Tyr(PO₃H₂)), alkylglycine,1-aminoindane-1-carboxylic acid, 2-aminoindane-2-carboxylic acid (Aic),4-amino-methylpyrrol-2-carboxylic acid (Py),4-amino-pyrrolidine-2-carboxylic acid (Abpc),2-aminotetraline-2-carboxylic acid (Atc), diaminoacetic acid (Gly(NH₂)),diaminobutyric acid (Dab), 1,3-dihydro-2H-isoinole-carboxylic acid(Disc), homocylcohexylalanine (hCha), homophenylalanine (hPhe or Hof),trans-3-phenyl-azetidine-2-carboxylic acid,4-phenyl-pyrrolidine-2-carboxylic acid,5-phenyl-pyrrolidine-2-carboxylic acid, 3-pyridylalanine (3-Pya),4-pyridylalanine (4-Pya), styrylalanine,tetrahydroisoquinoline-1-carboxylic acid (Tiq),1,2,3,4-tetrahydronorharmane-3-carboxylic acid (Tpi),β-(2-thienyl)-alanine (Tha).

“Peptides” are selected from dipeptides to decapeptides, preferred aredipeptides, tripeptides, tetrapeptides and pentapeptides. The aminoacids for the formation of the “peptides” can be selected from thoselisted above.

An “aza-amino acid” is defined as an amino acid where the chiral α-CHgroup is replaced by a nitrogen atom, whereas an “aza-peptide” isdefined as a peptide, in which the chiral α-CH group of one or moreamino acid residues in the peptide chain is replaced by a nitrogen atom.

Other amino acid substitutions for those encoded in the genetic code canalso be included in peptide compounds within the scope of the inventionand can be classified within this general scheme. Proteinogenic aminoacids are defined as natural protein-derived α-amino acids.Non-proteinogenic amino acids are defined as all other amino acids,which are not building blocks of common natural proteins. “Peptidemimetics” per se are known to a person skilled in the art. They arepreferably defined as compounds which have a secondary structure like apeptide and optionally further structural characteristics; their mode ofaction is largely similar or identical to the mode of action of thenative peptide; however, their activity (e.g. as an antagonist orinhibitor) can be modified as compared with the native peptide,especially vis à vis receptors or enzymes. Moreover, they can imitatethe effect of the native peptide (agonist). Examples of peptide mimeticsare scaffold mimetics, non-peptidic mimetics, peptides, peptide nucleicacids, oligopyrrolinones, vinylogpeptides and oligocarbamates. For thedefinitions of these peptide mimetics see Lexikon der Chemie, SpektrumAkademischer Verlag Heidelberg, Berlin, 1999.

The aim for using these mimetic structures is increasing the activity,increasing the selectivity to decrease side effects, protect thecompound against enzymatic degradation for prolongation of the effect.

Stereoisomers:

All possible stereoisomers of the claimed compounds are included in thepresent invention.

Where the compounds according to this invention have at least one chiralcenter, they may accordingly exist as enantiomers. Where the compoundspossess two or more chiral centers, they may additionally exist asdiastereomers. It is to be understood that all such isomers and mixturesthereof are encompassed within the scope of the present invention.

Preparation and Isolation of Stereoisomers:

Where the processes for the preparation of the compounds according tothe invention give rise to a mixture of stereoisomers, these isomers maybe separated by conventional techniques such as preparativechromatography. The compounds may be prepared in racemic form, orindividual enantiomers may be prepared either by enantiospecificsynthesis or by resolution. The compounds may, for example, be resolvedinto their components enantiomers by standard techniques, such as theformation of diastereomeric pairs by salt formation with an opticallyactive acid, such as (−)-di-p-toluoyl-d-tartaric acid and/or(+)-di-p-toluoyl-l-tartaric acid followed by fractional crystallizationand regeneration of the free base. The compounds may also resolved byformation of diastereomeric esters or amides, followed bychromatographic separation and removal of the chiral auxiliary.Alternatively, the compounds may be resolved using a chiral HPLC column.

Pharmaceutically Acceptable Salts:

In view of the close relationship between the free compounds and thecompounds in the form of their salts or solvates, whenever a compound isreferred to in this context, a corresponding salt or solvate is alsointended, provided such is possible or appropriate under thecircumstances.

Salts and solvates of the compounds of formula (1) and physiologicallyfunctional derivatives thereof which are suitable for use in medicineare those wherein the counter-ion or associated solvent ispharmaceutically acceptable. However, salts and solvates havingnon-pharmaceutically acceptable counter-ions or associated solvents arewithin the scope of the present invention, for example, for use asintermediates in the preparation of other compounds and theirpharmaceutically acceptable salts and solvates.

Suitable salts according to the invention include those formed with bothorganic and inorganic acids or bases. Pharmaceutically acceptable acidaddition salts include those formed from hydrochloric, hydrobromic,sulphuric, nitric, citric, tartaric, phosphoric, lactic, pyruvic,acetic, trifluoroacetic, triphenylacetic, sulphamic, sulphanilic,succinic, oxalic, fumaric, maleic, malic, mandelic, glutamic, aspartic,oxaloacetic, methanesulphonic, ethanesulphonic, arylsulphonic (forexample p-toluenesulphonic, benzenesulphonic, naphthalenesulphonic ornaphthalenedisulphonic), salicylic, glutaric, gluconic, tricarballylic,cinnamic, substituted cinnamic (for example, phenyl, methyl, methoxy orhalo substituted cinnamic, including 4-methyl and 4-methoxycinnamicacid), ascorbic, oleic, naphthoic, hydroxynaphthoic (for example 1- or3-hydroxy-2-naphthoic), naphthaleneacrylic (for examplenaphthalene-2-acrylic), benzoic, 4-methoxybenzoic, 2- or4-hydroxybenzoic, 4-chlorobenzoic, 4-phenylbenzoic, benzeneacrylic (forexample 1,4-benzenediacrylic), isethionic acids, perchloric, propionic,glycolic, hydroxyethanesulfonic, pamoic, cyclohexanesulfamic, salicylic,saccharinic and trifluoroacetic acid. Pharmaceutically acceptable basesalts include ammonium salts, alkali metal salts such as those of sodiumand potassium, alkaline earth metal salts such as those of calcium andmagnesium and salts with organic bases such as dicyclohexylamine andN-methyl-D-glucamine.

All pharmaceutically acceptable acid addition salt forms of thecompounds of the present invention are intended to be embraced by thescope of this invention.

Examples of solvates include hydrates.

Polymorph Crystal Forms:

Furthermore, some of the crystalline forms of the compounds may exist aspolymorphs and as such are intended to be included in the presentinvention. In addition, some of the compounds may form solvates withwater (i.e. hydrates) or common organic solvents, and such solvates arealso intended to be encompassed within the scope of this invention. Thecompounds, including their salts, can also be obtained in the form oftheir hydrates, or include other solvents used for theircrystallization.

Prodrugs:

The present invention further includes within its scope prodrugs of thecompounds of this invention. In general, such prodrugs will befunctional derivatives of the compounds which are readily convertible invivo into the desired therapeutically active compound. Thus, in thesecases, the methods of treatment of the present invention, the term“administering” shall encompass the treatment of the various disordersdescribed with prodrug versions of one or more of the claimed compounds,but which converts to the above specified compound in vivo afteradministration to the subject. Conventional procedures for the selectionand preparation of suitable prodrug derivatives are described, forexample, in “Design of Prodrugs”, ed. H. Bundgaard, Elsevier, 1985 andthe patent applications DE 198 28 113, DE 198 28 114, WO 99/67228 and WO99/67279 which are fully incorporated herein by reference.

Protective Groups:

During any of the processes for preparation of the compounds of thepresent invention, it may be necessary and/or desirable to protectsensitive or reactive groups on any of the molecules concerned. This maybe achieved by means of conventional protecting groups, such as thosedescribed in Protective Groups in Organic Chemistry, ed. J. F. W.McOmie, Plenum Press, 1973; and T. W. Greene & P. G. M. Wuts, ProtectiveGroups in Organic Synthesis, John Wiley & Sons, 1991, fully incorporatedherein by reference. The protecting groups may be removed at aconvenient subsequent stage using methods known from the art.

As used herein, the term “composition” is intended to encompass aproduct comprising the claimed compounds in the therapeuticallyeffective amounts, as well as any product which results, directly orindirectly, from combinations of the claimed compounds.

Carriers and Additives for Galenic Formulations:

Thus, for liquid oral preparations, such as for example, suspensions,elixirs and solutions, suitable carriers and additives mayadvantageously include water, glycols, oils, alcohols, flavoring agents,preservatives, coloring agents and the like; for solid oral preparationssuch as, for example, powders, capsules, gelcaps and tablets, suitablecarriers and additives include starches, sugars, diluents, granulatingagents, lubricants, binders, disintegrating agents and the like.

Carriers, which can be added to the mixture, include necessary and inertpharmaceutical excipients, including, but not limited to, suitablebinders, suspending agents, lubricants, flavorants, sweeteners,preservatives, coatings, disintegrating agents, dyes and coloringagents.

Soluble polymers as targetable drug carriers can includepolyvinylpyrrolidone, pyran copolymer,polyhydroxypropylmethacrylamidephenol,polyhydroxyethylaspartamidephenol, or polyethyleneoxidepolyllysinesubstituted with palmitoyl residue. Furthermore, the compounds of thepresent invention may be coupled to a class of biodegradable polymersuseful in achieving controlled release of a drug, for example, polyacticacid, polyepsilon caprolactone, polyhydroxy butyeric acid,polyorthoesters, polyacetals, polydihydropyrans, polycyanoacrylates andcross-linked or amphipathic block copolymers of hydrogels.

Suitable binders include, without limitation, starch, gelatin, naturalsugars such as glucose or betalactose, corn sweeteners, natural andsynthetic gums such as acacia, tragacanth or sodium oleate, sodiumstearate, magnesium stearate, sodium benzoate, sodium acetate, sodiumchloride and the like.

Disintegrators include, without limitation, starch, methyl cellulose,agar, bentonite, xanthan gum and the like.

Peptide Sequences

The peptides mentioned and used herein have the following sequences:

Aβ (1-42): Asp-Ala-Glu-Phe-Arg-His-Asp-Ser-Gly-Tyr-Glu-Val-His-His-Gln-Lys-Leu-Val-Phe-Phe-Ala-Glu-Asp-Val-Gly-Ser-Asn-Lys-Gly-Ala-Ile-Ile-Gly-Leu-Met-Val- Gly-Gly-Val-Val-Ile-AlaAβ (1-40): Asp-Ala-Glu-Phe-Arg-His-Asp-Ser-Gly-Tyr-Glu-Val-His-His-Gln-Lys-Leu-Val-Phe-Phe-Ala-Glu-Asp-Val-Gly-Ser-Asn-Lys-Gly-Ala-Ile-Ile-Gly-Leu-Met-Val- Gly-Gly-Val-ValAβ (3-42): Glu-Phe-Arg-His-Asp-Ser-Gly-Tyr-Glu-Val-His-His-Gln-Lys-Leu-Val-Phe-Phe-Ala-Glu-Asp-Val-Gly-Ser-Asn-Lys-Gly-Ala-Ile-Ile-Gly-Leu-Met-Val-Gly-Gly- Val-Val-Ile-AlaAβ (3-40): Glu-Phe-Arg-His-Asp-Ser-Gly-Tyr-Glu-Val-His-His-Gln-Lys-Leu-Val-Phe-Phe-Ala-Glu-Asp-Val-Gly-Ser-Asn-Lys-Gly-Ala-Ile-Ile-Gly-Leu-Met-Val-Gly-Gly- Val-Val Aβ (1-11)a:Asp-Ala-Glu-Phe-Arg-His-Asp-Ser-Gly-Tyr-Glu-NH₂ Aβ (3-11)a:Glu-Phe-Arg-His-Asp-Ser-Gly-Tyr-Glu-NH₂ Aβ (1-21)a:Asp-Ala-Glu-Phe-Arg-His-Asp-Ser-Gly-Tyr-Glu-Val-His-His-Gln-Lys-Leu-Val-Phe-Phe-Ala-NH₂ Aβ (3-21)a:Glu-Phe-Arg-His-Asp-Ser-Gly-Tyr-Glu-Val-His-His-Gln-Lys-Leu-Val-Phe-Phe-Ala-NH₂ Gln³-Aβ (3-40):Gln-Phe-Arg-His-Asp-Ser-Gly-Tyr-Glu-Val-His-His-Gln-Lys-Leu-Val-Phe-Phe-Ala-Glu-Asp-Val-Gly-Ser-Asn-Lys-Gly-Ala-Ile-Ile-Gly-Leu-Met-Val-Gly-Gly- Val-ValGln³-Aβ (3-21)a: Gln-Phe-Arg-His-Asp-Ser-Gly-Tyr-Glu-Val-His-His-Gln-Lys-Leu-Val-Phe-Phe-Ala-NH₂ Gln³-Aβ (1-11)a:Asp-Ala-Gln-Phe-Arg-His-Asp-Ser-Gly-Tyr-Glu-NH₂ Gln³-Aβ (3-11)a:Gln-Phe-Arg-His-Asp-Ser-Gly-Tyr-Glu-NH₂

SUMMARY OF THE INVENTION

The present invention provides compounds that act as inhibitors ofglutaminyl cyclase (QC, EC 2.3.2.5).

Physiological substrates of QC in mammals are, e.g. [Glu³] amyloidβ-protein (3-40/42), [Gln³] amyloid β-protein (3-40/42), Gastrin,Neurotensin, FPP, CCL 2, CCL 7, CCL 8, CCL 16, CCL 18, Fractalkine,Orexin A, [Gln³]-glucagon(3-29) and [Gln⁵]-substance P(5-11). Thecompounds according to the present invention and pharmaceuticalcompositions comprising at least one compound according to the presentinvention are useful for the treatment of conditions that can be treatedby modulation of QC activity.

By administering inhibitors of QC (EC) activity to a mammal it ispossible to prevent or alleviate or treat neuronal disorders(Alzheimer's disease, Down Syndrome, Parkinson disease, ChoreaHuntington, pathogenic psychotic conditions, schizophrenia, impairedfood intake, sleep-wakefulness, impaired homeostatic regulation ofenergy metabolism, impaired autonomic function, impaired hormonalbalance, impaired regulation, body fluids, hypertension, fever, sleepdysregulation, anorexia, anxiety related disorders including depression,seizures including epilepsy, drug withdrawal and alcoholism,neurodegenerative disorders including cognitive dysfunction anddementia).

Furthermore, by administration of a compound according to the presentinvention to a mammal it can be possible to stimulate the proliferationof myeloid progenitor cells.

In addition, the administration of a QC inhibitor according to thepresent invention can lead to suppression of male fertility.

In a preferred embodiment, the present invention provides the use ofinhibitors of QC (EC) activity in combination with other agents,especially for the treatment of neuronal disorders.

DETAILED DESCRIPTION OF THE INVENTION

The present invention provides novel inhibitors of QC (EC) of theformula 1,

wherein:A is either:

-   -   an alkyl chain, alkenyl chain or alkynyl chain;        or A is a group selected from:

-   -   wherein:    -   R⁶, R⁷, R⁸, R⁹ and R¹⁰ are independently H or an alkyl chain,        alkenyl chain, alkynyl chain, cycloalkyl, a carbocycle, aryl,        heteroaryl, or a heterocycle;    -   n and n¹ are independently 1-5;    -   m is 1-5;    -   o is 0-4;        and B is a group selected from (VI)-(XIV):

-   -   wherein:    -   D and E independently represent an alkyl chain, alkenyl chain,        alkynyl chain, a cycloalkyl, carbocycle, aryl, -alkylaryl,        heteroaryl, -alkylheteroaryl, acyl or a heterocycle.    -   X represents CR²⁰R²¹, O, S, NR¹⁹, with the proviso for        formulas (VIII) and (IX) that, if Z=CH, X is O or S;    -   R¹⁹ is selected from the group consisting of H, alkyl,        cycloalkyl, aryl, heteroaryl, -oxyalkyl, -oxyaryl, carbonyl,        amido, hydroxy, NO₂, NH₂, CN;    -   R²⁰ and R²¹ are independently selected from H, alkyl,        cycloalkyl, heterocycle, aryl, heteroaryl, -oxyalkyl, -oxyaryl,        carbonyl, amido, NO₂, NH₂, CN, CF₃;    -   X¹, X² and X³ are independently O or S provided that X² and X³        are not both O;    -   Y is O or S, with the proviso that Y may not be 0, when the        carbocycle formed by R¹⁷ and R¹⁸ has 3 members in the ring;    -   Z is CH or N;    -   R¹¹, R¹², R¹³ and R¹⁴ can be independently selected from H, an        alkyl chain, an alkenyl chain, an alkynyl chain, cycloalkyl,        carbocycle, aryl, heteroaryl, a heterocycle, halogen, alkoxy-,        -thioalkyl, carboxyl, carboxylic acid ester, carbonyl,        carbamide, carbimide, thiocarbamide or thiocarbonyl, NH₂, NO₂;    -   R¹⁵ and R¹⁶ are independently of each other H or a branched or        unbranched alkyl chain, or a branched or unbranched alkenyl        chain;    -   R¹⁷ and R¹⁸ are independently selected from H or an alkyl chain,        alkenyl chain, a alkynyl chain, a carbocycle, aryl, heteroaryl,        heteroalkyl or can be connected to form a carbocycle with up to        6 ring atoms;    -   n is 0 or 1;        with the proviso that the following compounds:

are excluded from formula 1.

When A is selected from an alkyl chain, alkenyl chain or alkynyl chain,preferably A is a C₁-C₇ alkyl chain, C₁-C₇ alkenyl chain or a C₁-C₇alkynyl chain. In one embodiment of the invention A is an unbranchedC₂₋₅ alkyl chain, in particular an unbranched C₃₋₄ alkyl chain,especially an unbranched C₃ alkyl chain. In a second embodiment of theinvention A represents a C₃ alkyl chain which is substituted at the 2position by one (i.e. in S or R configuration) or two methyl groups.

When A is selected from the formulae (I) to (V), preferably A isselected from groups (I) to (IV). In one embodiment of the invention Arepresents a group of formula (IV), wherein n¹ are each equal to 1 andm=1-4, especially m=1. In a second embodiment of the invention Arepresents a group of formula (I), (II) or (III), wherein n and n¹ areeach equal to 1 and R⁶, R⁷, R⁸, R⁹ and R¹⁰ represent H.

Preferably R⁶, R⁷, R⁸, R⁹ and R¹⁰ represent H or methyl.

In one embodiment of the invention the group B is chosen from (VI),(VIa), (VIb), (VII), (X), (XI), (XII), (XIII) and (XIV). In a secondembodiment of the invention group B represents formula (VI). In a thirdembodiment of the invention group B represents formula (VIa). In afourth embodiment of the invention group B represents formula (VIb). Ina fifth embodiment of the invention group B represents formula (VII). Ina sixth embodiment of the invention group B represents formula (X). In aseventh embodiment of the invention group B represents formula (XI). Inan eighth embodiment of the invention group B represents formula (XII).In another embodiment of the invention group B represents formula(XIII). In a further embodiment of the invention group B representsformula (XIV). In a preferred embodiment of the invention B represents agroup of formula (VI) or (VII).

When B represents a group (IX) suitably A does not represent alkynyl.

Preferably D and E independently represent benzyl, aryl, heteroaryl or aheterocycle.

In one embodiment of the invention D and E represent aryl, in particularphenyl or napthyl, especially substituted phenyl. Preferred substituentgroups when D represents phenyl include alkoxy-, -thioalkyl, halogen, ora carboxylic acid alkyl or aryl ester. Also preferred are fluoro,chloro, bromo, iodo, trifluoromethyl, trifluoromethoxy, methoxy, ethoxy,benzyloxy, cyano, acetyl, dimethyl amino, methylsulphanyl, nitro,oxazolyl, pyrazolyl, isopropyl, ethyl and methoxycarbonyl. Where aphenyl group is mono-substituted it is preferred that substitution is inthe 4-position. Other suitable aryl groups which D and E may representinclude dihydrobenzodioxine, benzodioxole, benzodithioledihydrobenzodithiine, benzooxathiole and dihydrobenzooxathiine. Aparticularly preferred group which D or E may represent is3,4-(dimethoxy)-phenyl,

Preferably R²⁰ and R²¹ represent NO₂, CN, CF₃ or, if R²⁰ is H, R²¹ isNO₂, CN, CF₃, or, if R²¹ is H, R²⁰ is NO₂, CN, CF₃.

In one embodiment, X or Y is S, O or NR¹. Preferably X or Y is S.

Preferably Z represents N.

In a preferred embodiment, R¹¹ and R¹⁴ are H.

In a further preferred embodiment, R¹² and R¹³ are independentlyselected from oxyalkyl or thioalkyl, halogen, or carboxylic acid alkylester or phenyl.

In a preferred embodiment, at least one of R¹⁵ and R¹⁶ is H, morepreferably, R¹⁵ and R¹⁶ are both H.

In a preferred embodiment, one of R¹⁷ and R¹⁸ is H and the other is Me.Also preferred are compounds wherein one of R¹⁷ and R¹⁸ is H and theother is phenyl. Additionally preferred are compounds where R¹⁷ and R¹⁸form a carbocycle with up to 6 members in the ring atoms.

Preferred compounds include those defined by Examples 13, 119 and 125below.

The present invention provides compounds of formula 1 for use as apharmaceutical. with the proviso that the compounds:

are excluded from formula 1.

The compound (a) of the proviso above is disclosed as compound 7 inGanellin et al (1995) J Med Chem 38(17) 3342-3350. This paper disclosessaid compound as a weak inhibitor of the histamine H3 receptor.

The compound of proviso (b) is disclosed as compound 7 in Venkatachalamet al (2001) Bioorganic Med Chem Lett 11, 523-528. This discloses saidcompound as an HIV1 reverse transcriptase inhibitor.

The compound of proviso (c) is disclosed as compound 19b in Moon et al(1991) J Med Chem 34, 2314-2327. This paper discloses said compound as acholinergic agonist with potential use in the treatment of Alzheimer'sdisease.

The compounds of proviso (d) are disclosed as compounds 99, 100 and102-103 in Wright et al (1986) J Med Chem 29, 523-530. This paperdiscloses said compounds as thromoxane synthetase inhibitors.

Certain compounds which would be embraced by formula 1 if it were notfor the proviso “provided that X² and X³ are not both O” are disclosedin Wright et al (1987) J Med Chem 30, 2277-2283 as thromboxanesynthetase inhibitors.

Certain compounds which would be embraced by formula 1 if it were notfor the proviso “that Y may not be 0, when the carbocycle formed by R¹⁷and R¹⁸ has 3 members in the ring” are disclosed in EP 0 117 462 A2 asthromboxane synthetase inhibitors.

Furthermore, the present invention provides the use of inhibitors of QCof the formula I, without the proviso excluding compounds (a)-(d) or theproviso that X² and X³ are not both O or the proviso that Y may not beO, when the carbocycle formed by R¹⁷ and R¹⁸ has 3 members in the ring,for the preparation of a medicament for the treatment of diseasesselected from the group consisting of Alzheimer's disease, DownSyndrome, Parkinson disease, Chorea Huntington, pathogenic psychoticconditions, schizophrenia, impaired food intake, sleep-wakefulness,impaired homeostatic regulation of energy metabolism, impaired autonomicfunction, impaired hormonal balance, impaired regulation, body fluids,hypertension, fever, sleep dysregulation, anorexia, anxiety relateddisorders including depression, seizures including epilepsy, drugwithdrawal and alcoholism, neurodegenerative disorders includingcognitive dysfunction and dementia.

The present invention also provides inhibitors of QC of the formula 1,without the proviso excluding compounds (a)-(d) or the proviso that X²and X³ are not both O or the proviso that Y may not be O, when thecarbocycle formed by R¹⁷ and R¹⁸ has 3 members in the ring, for use inthe treatment of diseases selected from the group consisting ofAlzheimer's disease, Down Syndrome, Parkinson disease, ChoreaHuntington, pathogenic psychotic conditions, schizophrenia, impairedfood intake, sleep-wakefulness, impaired homeostatic regulation ofenergy metabolism, impaired autonomic function, impaired hormonalbalance, impaired regulation, body fluids, hypertension, fever, sleepdysregulation, anorexia, anxiety related disorders including depression,seizures including epilepsy, drug withdrawal and alcoholism,neurodegenerative disorders including cognitive dysfunction anddementia.

The present invention also provides a method of treatment for a diseaseselected from the group consisting of Alzheimer's disease, DownSyndrome, Parkinson disease, Chorea Huntington, pathogenic psychoticconditions, schizophrenia, impaired food intake, sleep-wakefulness,impaired homeostatic regulation of energy metabolism, impaired autonomicfunction, impaired hormonal balance, impaired regulation, body fluids,hypertension, fever, sleep dysregulation, anorexia, anxiety relateddisorders including depression, seizures including epilepsy, drugwithdrawal and alcoholism, neurodegenerative disorders includingcognitive dysfunction and dementia, comprising the administration of atherapeutically active amount of at least one compound of formula 1,without the proviso excluding compounds (a)-(d) or the proviso that X²and X³ are not both O or the proviso that Y may not be O, when thecarbocycle formed by R¹⁷ and R¹⁸ has 3 members in the ring, to a mammal,preferably a human.

Most preferably, the present invention provides a method of treatmentand corresponding uses for a disease selected from the group consistingof Alzheimer's disease, Down Syndrome, Parkinson disease and ChoreaHuntington, comprising the administration of a therapeutically activeamount of at least one compound of formula 1, without the provisoexcluding compounds (a)-(d), to a mammal, preferably a human.

Suitably in the above mentioned methods and uses the compound is not thecompound of proviso (c).

A further compound, that of formula 1* shown below, is also a novelinhibitor of QC:

The compound of formula 1* may be employed in the methods and usesaccording to the invention in an analogous manner to the compounds offormula 1 described above.

In a further embodiment, the present invention provides novel inhibitorsof QC (EC) of the formula 1a,

wherein R is defined in examples 1 to 53.

Res. ESI-MS Act. IC₅₀ K_(i) Example R (M + H) (%) (μM) (μM) 1 Methyl199.3 4.3 13 2 tert-Butyl 241.4 60.7 14.7 3 Benzyl 275.4 60.9 5.67 4Phenyl 261.4 42.3 4.4 5 4-(fluoro)-phenyl 279.35 42.0 4.73 64-(chloro)-phenyl 295.80 1.2 7 4-(ethyl)-phenyl 289.41 28.7 2.78 84-(trifluoromethyl)- 329.4 38.5 3.93 phenyl 9 4-(methoxy-carbonyl)-319.4 1.19 Phenyl 10 4-(acetyl)-phenyl 303.4 17.0 1.70 114-(methoxy)-phenyl 291.4 9.7 0.70 12 bicyclo[2.2.1]hept- 277.5 16.05-en-2-yl 13 3,4-(dimethoxy)-phenyl 321.5 0.7 0.22 0.06 142,4-(dimethoxy)-phenyl 321.5 2.2 0.57 15 3,5-(dimethoxy)-phenyl 321.52.86 0.75 16 2-(methoxy-carbonyl)- 319.4 Phenyl 17 4-(oxazol-5-y)-phenyl328.5 3.64 0.86 18 4-(pyrazol-1-yl)-phenyl 327.4 19 4-(isopropyl)-phenyl303.5 8.7 20 4-(piperidine-1-sulfonyl)- 408.6 8.5 2.27 Phenyl 214-(morpholin-4-yl)-phenyl 346.5 9.0 22 4-(cyano)-phenyl 286.4 9.0 2.8923 2,3-dihydro-benzo[1,4] 319.4 4.17 1.12 dioxin-6-yl 24benzo[1,3]dioxol-5-yl 305.4 16.7 5.66 25 3,4,5(trimethoxy)-phenyl 351.51.7 0.34 26 3-(methoxy)-phenyl 291.4 6.8 1.86 27 4-(ethoxy)-phenyl 305.57.2 0.89 28 4-(benzyloxy)-phenyl 367.5 0.98 29 4-(methoxy)-benzyl 305.53.93 30 3,4-(dimethoxy)-benzyl 335.5 1.55 31 2-(methoxy-carbonyl)- 325.5thiophene-3-yl 32 3-(ethoxy-carbonyl)- 392.6 4,5,6,7-tetrahydrobenzo[b]thio- phene2-yl 33 2-(methoxy-carbonyl)-4- 339.5(methyl)-thiophene-3-yl 34 Benzo[c][1,2,5] 319.5 thiazol-4-yl 35Benzo[c][1,2,5] 319.5 4.4 1.37 thiazol-5-yl 36 5-(methyl)-3-(phenyl)-342.5 isooxazol-4-yl 37 3,5-(dimethyl)-isooxazol- 280.4 4-yl 384-(iodo)-phenyl 387.3 23.5 2.12 39 4-(bromo)-phenyl 340.3 2.52 404-(methyl)-phenyl 275.4 31.3 2.14 41 Naphthalen-1-yl 311.5 26.7 2.79 424-(nitro)-phenyl 306.4 31.1 2.68 43 Butyl 241.4 53.8 14.0 44 Cyclooctyl295.5 33.1 9.1 45 Furan-2-ylmethyl 265.4 61.4 10.0 46Tetrahydrofuran-2-ylmethyl 269.4 46.0 12.8 47 Benzo[1,3]dioxol-5- 319.442.7 6.1 ylmethyl 48 2-(morpholin-4-yl)-ethyl 298.5 55.0 13.3 494-(methylsulfanyl)-phenyl 307.5 19.1 1.66 50 4-(dimethylamino)-phenyl304.5 2.03 51 4-(trifluoromethoxy)-phenyl 345.4 14.2 52 Benzoyl 288.3 53Pyridin-4-yl 261.1

In a further embodiment, the present invention provides novel inhibitorsof QC (EC) of the formula 1b,

wherein R¹ and R² are defined in examples 54 to 95.

ESI-MS Res. Act. K_(i) Example R¹ R² (M + H) (%) (μM) 54 Cyano Methyl207.3 1.5 55 Cyano 3,4-(dimethoxy)-phenyl 329.4 1.36 56 Cyano2,4-(dimethoxy)-phenyl 329.4 57 Cyano 3,5-(dimethoxy)-phenyl 329.4 0.9158 Cyano 2,3-dihydrobenzo[b] 327.4 0.64 [1,4]dioxin-7-yl 59 CyanoBenzo[d][1,3]dioxol-6-yl 313.4 0.73 60 Cyano 3,4,5-(trimethoxy)-phenyl359.4 0.88 61 Cyano 3-(methoxy)-phenyl 299.4 62 Cyano 4-(ethoxy)-phenyl313.4 63 Cyano 4-(benzyloxy)-phenyl 375.5 64 Cyano Phenyl 269.4 1.02 65Cyano 4-(methoxy)-phenyl 299.4 0.70 66 Cyano 4-(acetyl)-phenyl 311.4 67Cyano 4-(nitro)-phenyl 314.4 68 Cyano Benzyl 283.4 22.5 8.17 69 CyanoNaphthalen-1-yl 319.4 70 Cyano 4-(fluoro)-phenyl 387.3 71 Cyano4-(iodo)-phenyl 395.3 72 Cyano 4-(bromo)-phenyl 348.3 73 CyanoCyclooctyl 289.4 74 Cyano tert-butyl 249.3 75 Cyano 4-(methyl)-phenyl283.3 1.34 76 Cyano 4-(methylthio)-phenyl 315.5 77 Cyano4-(ethyl)-phenyl 297.4 78 Cyano 4-(dimethylamino)-phenyl 312.4 79 CyanoButyl 249.4 80 Cyano Trityl 435.6 81 Cyano (Benzo[d][1,3]-dioxol- 327.41.53 6yl)methyl 82 Cyano (tetrahydrofuran-2yl)methyl 277.4 83 Cyano4-(trifluoromethyl)-phenyl 334.4 84 Cyano (furan-2-yl)methyl 273.4 85Cyano 2-(morpholin-4-yl)-ethyl 306.4 86 Cyano 4-(oxazol-5yl)-phenyl336.4 87 Cyano Pyridin-3-yl 270.4 88 Cyano 4-(cyano)-phenyl 294.4 89Cyano 4-(trifluoromethoxy)-phenyl 353.4 90 Cyano4-(piperidinosulfonyl)-phenyl 416.6 91 Cyano 4-(1H-pyrazol-1-yl)phenyl335.4 92 H 3,4-(dimethoxy)-phenyl 304.4 204.5 93 Methyl3,4-(dimethoxy)-phenyl 318.4 3.62 94 Cyano 2,3,4-(trimethoxy)-phenyl358.1 95 Cyano Cycloheptyl 288.2

In a further embodiment, the present invention provides novel inhibitorsof QC (EC) of the formula 1c,

wherein R³ is defined in examples 96 to 102.

Res. ESI-MS Act. IC₅₀ K_(i) Example R³ (M + H) (%) (μM) (μM) 96 Ethyl197.3 19.2 97 6-fluoro-4H-benzo[d] 321.4 19.0 12.0 [1,3]dioxin-8-yl 983-(cylopentyloxy)-4- 359.4 2.87 0.62 (methoxy)-phenyl 994-(heptyloxy)-phenyl 359.5 5.6 9.9 100 3,4-dihydro-2H-benzo[b] 317.4[1,4]dioxepin-7-yl 101 4-(butoxy)-phenyl 317.4 1023,4-(dimethoxy)-phenyl 305.4 0.46

In a further embodiment, the present invention provides novel inhibitorsof QC (EC) of the formula 1d,

wherein the position on the ring is defined in examples 103 to 105.

Position of the Benzyl- ESI-MS Res. Act. K_(i) Example substitution (M +H) (%) (μM) 103 2 383.5 16.27 4.84 104 3 383.5 3.52 105 4 383.5 1.86

In a further embodiment, the present invention provides novel inhibitorsof QC (EC) of the formula 1e,

wherein R⁴ and R⁵ are defined in examples 106 to 109.

ESI-MS Res. Act. IC₅₀ K_(i) Example R⁴ R⁵ (M + H) (%) (μM) (μM) 106(S) HMethyl 335.5 0.76 107(R) Methyl H 335.5 0.35 108 Methyl Methyl 349.5 109—CH₂—CH₂— 347.5 7.85

In a further embodiment, the present invention provides novel inhibitorsof QC (EC) of the formula 1f,

wherein R⁶ is defined in examples 110 to 112.

ESI-MS Res. Act. IC₅₀ K_(i) Example R⁶ (M + H) (%) (μM) (μM) 110 H 259.43.00 111 Chloro 293.8 3.35 112 Methoxy 289.4 1.57

In a further embodiment, the present invention provides novel inhibitorsof QC (EC) of the formula 1g,

wherein R⁷, R⁸ and R⁹ are defined in examples 113 to 132.

ESI-MS Res. Act. K_(i) Example R⁷ R⁸ R⁹ (M + H) (%) (μM) 113 Phenyl H H260.4 4.62 114 Thiophen-2-yl H H 266.5 3.29 115(R) Phenyl Methyl H 274.521.2 7.34 116(S) Phenyl H Methyl 274.5 8.1 3.51 117 Phenyl H Ethyl 288.53.57 118 Phenyl H Phenyl 336.5 13.5 4.48 119 3,4-(dimethoxy)- H H 320.50.39 Phenyl 120 3,4-(dimethoxy)- Methyl Methyl 347.2 Phenyl 1214-(chloro)-phenyl —CH₂—CH₂—CH₂— 334.9 4.88 122 4-(chloro)-phenyl—CH₂—C₂H₄—CH₂— 349.0 7.3 123 4-(methoxy)-phenyl —Ch₂—C₃H₆—CH₂— 358.62.78 124 4-(methoxy)-phenyl —CH₂—CH₂— 316.5 0.39 125 3,4-(dimethoxy)-—CH₂—CH₂— 346.5 0.09 Phenyl 126 3,4,5-(trimethoxy)- —CH₂—CH₂— 376.6Phenyl 127 2,3,4-(trimethoxy)- —CH₂—CH₂— 376.6 Phenyl 1282-(methoxy)-phenyl —CH₂—CH₂— 316.5 129 3-(methoxy)-phenyl —CH₂—CH₂—316.5 130 2,3-(dimethoxy)- —CH₂—CH₂— 346.5 Phenyl 131 3,5-(dimethoxy)-—CH₂—CH₂— 346.5 Phenyl 132 2,5-(dimethoxy)- —CH₂—CH₂— 346.5 Phenyl

In a further embodiment, the present invention provides novel inhibitorsof QC (EC) of the formula 1h,

wherein n is defined in examples 133 to 135.

ESI-MS K_(i) Example N (M + H) (μM) 133 3 306.4 134 4 320.5 0.99 135 5334.5

In a further embodiment, the present invention provides novel inhibitorsof QC (EC) of the formula 1i,

wherein m is defined in examples 136 and 137.

ESI-MS Res. Act. K_(i) Example m (M + H) (%) (μM) 136 2 307.4 17.6 137 4335.5 2.19 0.55

Further novel inhibitors of QC (EC) are examples 138 to 141.

ESI-MS Res. Act. IC₅₀ K_(i) Example Structure (M + H) (%) (μM) (μM) 138

347.5 139

347.2 140

226.3 13.8 20.5 141

370.4

SYNTHESIS OF THE EXAMPLES

Analytical Conditions

ESI-Mass spectra were obtained with a SCIEX API 365 spectrometer (PerkinElmer). The ¹H-NMR (500 MHz) data was recorded on a BRUKER AC 500, usingDMSO-D₆ as solvent. Chemical shifts are expressed as parts per milliondownfield from tetramethylsilane. Splitting patterns have beendesignated as follows: s (singulet), d (doublet), dd (doublet ofdoublet), t (triplet), m (multiplet), and br (broad signal).

Detailed Synthesis Description EXAMPLES 1-12 AND 14-53

1H-imidazole-1-propanamine was reacted with the correspondingisothiocyanate in ethanol under reflux for 8 h. After that the solventwas removed and the remaining oil was dissolved in methylene chloride.The organic layer was washed twice with a saturated solution of NaHCO₃followed by NaHSO₄ and brine, dried then evaporated. The remaining solidwas re-crystallized from ethyl acetate, yielding the example thiourea inyields of 80-98%.

EXAMPLE 131-(3-(1H-imidazol-1-yl)propyl)-3-(3,4-dimethoxyphenyl)thiourea

4.0 mmol of 3,4-dimethoxyphenyl isothiocyanate and 4.0 mmol of3-(1H-imidazol-1-yl)alkyl-1-amine were dissolved in 10 mL of absoluteethanol. After stirring for 2 h under reflux, the solvent was evaporatedand the resulting solid was recrystallized from ethanol.

Yield: 0.66 g (51.3%); mp: 160.0-161.0° C.

¹H NMR 61.8-2.0 (m, 2H), 3.4-3.5 (m, 2H), 3.75 (s, 6H), 3.9-4.0 (m, 2H),6.7-6.8 (m, 1H), 6.9 (br m, 2H), 6.95 (s, 1H), 7.15 (s, 1H), 7.55 (br s,1H), 7.6 (s, 1H), 9.3 (s, 1H); MS m/z 321.2 (M+H), 253.3 (M—C₃H₃N₂.)

EXAMPLES 96-102

1H-imidazole-1-propanamine was reacted with the corresponding isocyanatein ethanol under reflux for 8 h. After that the solvent was removed andthe remaining oil was dissolved in methylene chloride. The organic layerwas washed twice with a saturated solution of NaHCO₃ followed by NaHSO₄and brine, dried then evaporated. The remaining solid wasre-crystallized from ethyl acetate, yielding the example urea in yieldsof 85-90%.

EXAMPLES 136, 137

The 1H-imidazole-1-alkylamines were prepared according to the literaturefrom c-brom-alkyl-phtalimides and imidazolium salt and. subsequenthydrazinolysis. The resulting products were transformed into thethioureas according to example 1-53 giving a 88% (example 136) and 95%(example 137) yield.

EXAMPLES 54-95

All examples were made from the corresponding thioureas by reacting withWater-soluble-carbodiimide (WSCD) and 1H-imidazole-1-propanamine in drydimethyl form-amide for 2 h at r.t. giving the trisubstituted guanidineswith yields from 40-87%.

EXAMPLES 103-105

Imidazole was reacted with the corresponding brommethylphenylcyanide inDMF, utilizing 1 equivalent of NaH for 3 h under rt., giving the1H-imidazole-1-methylphenylcyanides. The solvent was removed and theresulting oil was re-dissolved in dioxane. The cyanides were convertedin the corresponding amines using 1 equivalent of LiAlH₄. After adding asaturated solution of KHSO₄, dioxane was evaporated and the aqueouslayer was extracted by means of CHCl₃. The organic layer wasconcentrated in vacuo and the amine was converted in the correspondingthioureas according to example 1-53 giving a 78% (example 103) and 65%(example 104) and 81% (example 105) yield.

EXAMPLES 106-109

Starting from the correspondingmethansulfonate-2-methylpropyl-phthalimides the amines were synthesizedas described for the amines in example 136-137. The resulting productswere transformed into the thioureas according to example 1-53 givingexample 106-109 in total yields of 25-30%.

EXAMPLES 110-112

1H-imidazole-1-propanamine was reacted with the corresponding2-chlorobenzo[d]thiazole in toluol for 24 h at a temperature of 130° C.After removing the solvent and recristallization from methanol example110-112 was yielded in an amount of 55-65%.

EXAMPLES 113-118, 120-124 AND 126-132

1H-imidazole-1-propanamine was reacted with the corresponding 2-phenylacetic acid in dry dioxane by adding one equivalent of CAIBE andN-methylmorpholine at a temperature of 0° C. After 2 h the mixture wasallowed to warm to r.t. and the mixture was stirred for 12 h. Afterremoving the solvent the resulting oil was redissolved in methylenechloride and the organic layer was washed by means of an aqueoussolution of NaHCO₃ and water, dried and the solvent was evaporated. Theremaining oil was dissolved in dioxane adding Laweson's Reagent. Afterstirring for 12 h a saturated solution of NaHCO₃ was added. Dioxane wasevaporated and the aqueous layer was extracted by means of ethylacetate. The organic layer was separated, dried and the solvent wasevaporated. The remaining solid was crystallized from acetylacetate/ether, giving 113-118, 120-124 and 126-132 with total yields of62-85%.

EXAMPLE 119N-(3-(1H-imidazol-1-yl)propyl)-2-(3,4-dimethoxyphenyl)ethanethioamide

A mixture of 4.0 mmol triethylamine and 4.0 mmol of3-(1H-imidazol-1-yl)alkyl-1-amine 20 mL of dioxane was added drop wiseto an ice cooled, stirred solution of 4.0 mmol of2-(3,4-dimethoxyphenyl)acetyl chloride in 30 mL of dioxane. The mixturewas allowed to warm to r.t., and then stirred for 1 h. After removingthe solvent by reduced pressure, the residue was redissolved in 50 mL ofdichloromethane. The organic layer was washed by means of 30 mL ofsaturated aqueous solution of NaHCO₃, and water. The organic solutionwas dried, filtered, and the solvent was removed under reduced pressure.After redissolving in 50 mL of dry dioxane 2.2 mmol of Lawesson'sreagent was added, and the mixture was heated to 90° C. and stirred for8 h. The solvent was removed by reduced pressure, and the residue wasredissolved in 50 mL of dichloromethane. The organic layer was washedthree times by means of a saturated aqueous solution of NaHCO₃, followedthree times by water, dried, filtered, and then the organic solvent wasremoved. The compound was purified by chromatography using acentrifugal-force-chromatography device, (Harrison Research Ltd.)utilizing silica plates of a layer thickness of 2 mm, and a CHCl₃/MeOHgradient as eluting system.

Yield: 0.14 g (10.6%); melting point: 148.0-150.0° C.

¹H NMR δ 2.0-2.15 (br m, 2H), 3.4-3.5 (m, 2H), 3.7 (s, 6H), 6.75-6.8 (m,2H), 4.1-4.2 (m, 2H), 6.8-6.9 (m, 2H), 6.95-7.0 (m, 1H), 7.4 (s, 1H),7.75-7.85 (br m, 1H), 8.6 (s, 1H), 10.2 (s, 1H); MS m/z 320.2 (M+H),252.2 (M—C₃H₃N₂.)

EXAMPLE 125N-(3-(1H-imidazol-1-yl)propyl)-1-(3,4-dimethoxyphenyl)cyclopropanecarbothioamide

11.06 mmol of 3,4-dimethoxyphenyl acetonitrile, 34.8 mmol of2-Bromo-1-chloroethanole and 1.16 mmol of triethylbenzylammoniumhydrochloride were dissolved in 10 mL of an aqueous solution of KOH(60%). The mixture was transferred into an ultrasonic bath andvigorously stirred for 3 h at room temperature. The resulting suspensionwas diluted with 40 mL of water and extracted three times by means of 20mL of dichloromethane. The combined organic layers where washed by meansof an aqueous solution of hydrochloric acid (1N), dried over Na₂SO₄ andthe solvent was removed under reduced pressure. The remaining oil waspurified by flash-chromatography using silica gel and ethylacetate/heptane as eluting system, resulting in 0.81 g (34.4%) of1-(3,4-dimethoxyphenyl)cyclopropanecarbonitrile 3.9 mmol of1-(3,4-dimethoxyphenyl)cyclopropanecarbonitrile and 11.2 mmol of KOHwere suspended in 80 mL of ethylene glycol. The mixture was stirred for12 h under reflux. Then 80 mL of water were added and the aqueous layerwas extracted two times with ether. After pH adjustment to a value ofpH=4-5 using HCl (1N) the aqueous layer was extracted three times bymeans of ether, then the combined organic layers were dried over Na₂SO₄and the solvent was removed, resulting in 0.81 g (93.5%) of1-(3,4-dimethoxyphenyl)cyclopropanecarboxylic acid.

3.44 mmol of 1-(3,4-dimethoxyphenyl)cyclopropanecarboxylic acid, 3.5mmol of N-Methyl morpholine, and 3.5 mmol of isobutyl chloroformiat weredissolved in dry tetrahydrofurane and stirred for 15 min at −15° C. Then3.5 mmol of 3-(1H-imidazol-1-yl)alkyl-1-amine was added and the mixturewas allowed to warm to 0° C. and was stirred for 12 h. The solvent wasremoved under reduced pressure and the remaining oil was redissolved inchloroform. Then the organic layer was washed two times by means of asaturated aqueous solution of NaHCO₃, then dried over Na₂SO₄ and thesolvent was removed. Purification was performed by means of centrifugalforced chromatography using a chromatotron® device (Harrison ResearchLtd.) utilizing silica plates of a layer thickness of 2 mm, and aCHCl₃/MeOH gradient as eluting system resulting in 0.671 g (59.3%) ofN-(3-(1H-imidazol-1-yl)propyl)-1-(3,4-dimethoxyphenyl)cyclopropane-carboxamide.

After redissolving in 30 mL of dry dioxane 1.43 mmol of Lawesson'sreagent were added, and the mixture was heated to 90° C. and stirred for8 h. The solvent was removed by reduced pressure, and the residue wasremains were dissolved in 50 mL of dichloromethane. The organic layerwas washed three times by means of a saturated aqueous solution ofNaHCO₃, followed three times by water, dried, filtered, and then theorganic solvent was removed. The compound was purified by chromatographyusing a centrifugal-force-chromatography device, (Harrison ResearchLtd.) utilizing silica plates of a layer thickness of 2 mm, and aCHCl₃/MeOH gradient as eluting system.

Yield: 0.33 g (46.2%); melting point: 127.0-127.5° C.

¹H NMR δ 1.1-1.2 (t, 2H), 1.55-1.6 (t, 2H), 2.0-2.1 (m, 2H), 3.5-3.6 (m,2H), 3.7-3.8 (s, 6H), 4.1-4.2 (t, 2H), 6.8-6.9 (m, 3H), 7.65 (s, 1H),7.75 (s, 1H), 8.8 (m, 1H), 9.05 (s, 1H; MS m/z 346.0 (M+H), 278.2(M—C₃H₃N₂), 177.1 (M—C₆H₈N₃S)

EXAMPLES 133-135

A mixture of 1 equivalent triethylamine and 3,4-dimethoxyaniline indioxane was added to an stirred solution of the correspondingω-bromoalkyl acidic chloride at a temperature of 0° C. The solution wasallowed to warm to r.t. and stirred for 2 h. The solvent was evaporated,and the remaining oil was redissolved in dichloromethane. The organiclayer was washed by means of water, dried, filtered, and the solvent wasremoved under reduced pressure.

Imidazole and sodium hydride were suspended in and the mixture wasstirred under inert conditions at r.t. for 3 h.ω-Bromo-N-(3,4-dimethoxy-phenyl)alkylamide was added and the mixture washeated to 100° C. and stirred for 8 h. After that, the solvent wasevaporated, hot toluene were added and the solution was filtered. Thenthe solvent was removed under reduced pressure. The transformation intothe thioamides was performed as described for example 113-132 by meansof Laweson's reagent, giving 133-135 in total yields of 13-20%.

The analytical data for further examples, which were synthesizedaccording to the general synthesis schemes described above, are asfollows:

EXAMPLE 1 1-(3-(1H-imidazo/1-yl)propy)-3-methylthiourea

melting point: 122-122.5° C.

¹H NMR δ 1.85-1.95 (m, 2H), 2.8 (s, 3H), 3.2-3.5 (br d, 2H), 3.8-3.9 (m,2H), 6.85 (d, 1H), 7.15 (d, 1H), 7.3-7.5 (br d, 2H), 7.65 (s, 1H); MSm/z 199.1 (M+H), 221.3 (M+Na), 131.0 (M—C₃H₃N₂.)

EXAMPLE 2 1-(3-(1H-imidazol-1-yl)propyl)-3-tert-butylthiourea

melting point: 147.0-147.5° C.

¹H NMR δ 1.3-1.4 (s, 9H), 1.85-1.95 (m, 2H), 3.5 (t, 2H), 3.8 (t, 2H),6.85 (d, 1H), 7.15 (d, 1H), 7.3-7.5 (br d, 2H), 7.65 (s, 1H); MS m/z241.1 (M+H), 173.1 (M—C₃H₃N₂.)

EXAMPLE 3 1-(3-(1H-imidazol-1-yl)propyl)-3-benzylthiourea

melting point: 127.0-128.0° C.

¹H NMR δ 1.85-1.95 (m, 2H), 3.2-3.5 (br d, 2H), 3.8-3.9 (m, 2H), 4.6 (s,2H), 6.8 (d, 1H), 7.15 (d, 1H), 7.19-7.35 (m, 5H), 7.5-7.6 (br d, 2H),7.85 (s, 1H); MS m/z 275.3 (M+H), 207.1 (M—C₃H₃N₂.)

EXAMPLE 5 1-(3-(1H-imidazol-1-yl)propyl)-3-phenylthiourea

melting point: 166.5-167.0° C.

¹H NMR δ 1.95-2.05 (m, 2H), 3.3-3.5 (br d, 2H), 3.9-4.0 (m, 2H), 6.85(d, 1H), 7.05 (m, 1H) 7.15 (d, 1H), 7.25 (m, 2H), 7.35 (m, 2H), 7.6 (s,1H), 7.8 (br s, 1H), 9.5 (br s, 1H); MS m/z 261.1 (M+H), 193.2(M—C₃H₃N₂.)

EXAMPLE 6 1-(3-(1H-imidazol-1-yl)propyl)-3-(4-fluorophenyl)thiourea

melting point: 147.0-148.0° C.

¹H NMR δ 1.95-2.05 (m, 2H), 3.3-3.5 (br d, 2H), 3.9-4.05 (m, 2H), 6.85(d, 1H), 7.05-7.15 (m, 3H), 7.3-7.4 (m, 2H), 7.6 (s, 1H), 7.7-7.8 (br s,1H), 9.4 (br s, 1H); MS m/z 279.3 (M+H), 211.2 (M—C₃H₃N₂.)

EXAMPLE 7 1-(3-(1H-imidazol-1-yl)propyl)-3-(4-ethylphenyl)thiourea

melting point: 100.0-100.5° C.

¹H NMR δ 1.15-1.2 (t, 3H), 1.9-2.0 (m, 2H), 2.5-2.6 (m, 2H), 3.3-3.5 (brd, 2H), 3.9-4.05 (m, 2H), 6.85 (d, 1H), 7.1-7.2 (m, 3H), 7.25-7.3 (m,2H), 7.6 (s, 1H), 7.7-7.8 (br s, 1H), 9.4 (br s, 1H); MS m/z 289.3(M+H), 221.1 (M—C₃H₃N₂.)

EXAMPLE 81-(3-(1H-imidazol-1-yl)propyl)-3-(4-(trifluoromethyl)phenyl)thiourea

melting point: 154.5-155.0° C.

¹H NMR δ 1.9-2.1 (br m, 2H), 3.4-3.6 (br d, 2H), 3.95-4.1 (br m, 2H),6.85 (d, 1H), 7.2 (d, 1H), 7.6-7.8 (m, 5H), 8.2 (br s, 1H), 9.9 (br s,1H); MS m/z 329.3 (M+H), 261.2 (M—C₃H₃N₂.)

EXAMPLE 10 1-(3-(1H-imidazol-1-yl)propyl)-3-(4-acetylphenyl)thiourea

melting point: 170.0-171.0° C.

¹H NMR δ 1.9-2.1 (br m, 2H), 2.4-2.5 (s, 3H), 3.2-3.5 (br m, 2H),3.9-4.1 (m, 2H), 6.85 (d, 1H), 7.15 (d, 1H), 7.5-7.65 (br m, 3H),7.8-7.9 (m, 2H), 8.1 (m, 2H), 9.8 (br s, 1H); MS m/z 303.2 (M+H), 235.1(M—C₃H₃N₂.)

EXAMPLE 11 1-(3-(1H-imidazol-1-yl)propyl)-3-(4-methoxyphenyl)thiourea

melting point: 125.0-125.5° C.

¹H NMR δ 1.8-2.0 (br m, 2H), 3.2-3.5 (br m, 2H), 3.7 (s, 3H), 3.9-4.0(m, 2H), 6.7-6.9 (m, 3H), 7.1-7.2 (m, 3H), 7.5 (s, 1H), 7.6 (s, 1H), 9.2(s, 1H); MS m/z 291.1 (M+H), 223.2 (M—C₃H₃N₂.)

EXAMPLE 141-(3-(1H-imidazol-1-yl)propyl)-3-(2,4-dimethoxyphenyl)thiourea

melting point: 120.0-120.5° C.

¹H NMR δ 1.8-2.0 (br m, 2H), 3.4-3.5 (br m, 2H), 3.75 (s, 6H), 3.9-4.0(m, 2H), 6.5 (d, 1H), 6.6 (s, 1H), 6.9 (s, 1H), 7.15 (s, 1H), 7.3 (d,1H), 7.5 (br s, 1H), 7.6 (s, 1H), 9.75 (s, 1H); MS m/z 321.2 (M+H),253.3 (M—C₃H₃N₂.)

EXAMPLE 151-(3-(1H-imidazol-1-yl)propyl)-3-(3,5-dimethoxyphenyl)thiourea

melting point: 142.0-143.0° C.

¹H NMR δ 1.8-2.0 (br m, 2H), 3.4-3.5 (br m, 2H), 3.6 (s, 6H), 3.95-4.0(m, 2H), 6.25 (m, 1H), 6.6 (m, 2H), 6.9 (s, 1H), 7.2 (s, 1H), 7.6 (s,1H), 7.8 (s, 1H), 9.5 (s, 1H); MS m/z 321.2 (M+H), 253.3 (M—C₃H₃N₂.)

EXAMPLE 231-(3-(1H-imidazol-1-yl)propyl)-3-(2,3-dihydrobenzo[b][1,4]dioxin-7-yl)-thiourea

melting point: 103.0-103.5° C.

¹H NMR δ 1.9-2.0 (br m, 2H), 3.3-3.5 (br d, 2H), 3.9-4.0 (m, 2H),4.2-4.3 (m, 4H), 6.7 (m, 1H), 6.8-6.8 (m, 1H), 6.9 (m, 2H), 7.2 (s, 1H),7.6 (m, 2H), 9.3 (s, 1H); MS m/z 319.3 (M+H), 251.3 (M—C₃H₃N₂.)

EXAMPLE 241-(3-(1H-imidazol-1-yl)propyl)-3-(benzo[d][1,3]dioxol-6-yl)thiourea

melting point: 115.0-115.6° C.

¹H NMR δ 1.9-2.1 (br m, 2H), 3.4-3.5 (br d, 2H), 4.05-4.15 (m, 2H), 6.0(s, 2H), 6.7 (m, 1H), 6.8-6.85 (m, 1H), 6.95 (d, 1H), 7.25 (s, 1H), 7.45(s, 1H), 7.7 (br s, 1H), 8.5 (br s, 1H), 9.4 (br s, 1H); MS m/z 305.2(M+H), 237.2 (M—C₃H₃N₂.)

EXAMPLE 251-(3-(1H-imidazol-1-yl)propyl)-3-(3,4,5-trimethoxyphenyl)thiourea

melting point: 124.5-125.5° C.

¹H NMR δ 1.8-2.0 (m, 2H), 3.4-3.5 (br m, 2H), 3.6 (s, 3H), 3.7 (s, 6H),3.9-4.0 (m, 2H), 6.65 (m, 2H), 6.85 (s, 1H), 7.2 (s, 1H), 7.6 (s, 1H),7.7 (br s, 1H), 9.4 (s, 1H); MS m/z 351.3 (M+H), 283.2 (M—C₃H₃N₂.)

EXAMPLE 26 1-(3-(1H-imidazol-1-yl)propyl)-3-(3-methoxyphenyl)thiourea

melting point: 89.5-90.0° C.

¹H NMR δ 1.9-2.1 (br m, 2H), 3.4-3.5 (br m, 2H), 3.7 (s, 3H), 3.9-4.0(m, 2H), 6.6-6.7 (m, 1H), 6.8-6.9 (m, 2H), 7.1 (m, 2H), 7.15-7.25 (br m,1H), 7.6 (s, 1H), 7.8 (br s, 1H), 9.5 (s, 1H); MS m/z 291.1 (M+H), 223.2(M—C₃H₃N₂.)

EXAMPLE 27 1-(3-(1H-imidazol-1-yl)propyl)-3-(4-ethoxyphenyl)thiourea

melting point: 126.0-126.5° C.

¹H NMR δ 1.5 (br m, 3H), 1.9-2.0 (br m, 2H), 3.4-3.5 (br m, 2H), 3.9-4.0(br m, 4H), 6.8-6.9 (m, 2H), 6.95 (s, 1H), 7.15-7.2 (m, 2H), 7.25 (s,1H), 7.55-7.6 (br s, 1H), 7.8 (s, 1H), 9.3 (s, 1H); MS m/z 305.2 (M+H),237.2 (M—C₃H₃N₂.)

EXAMPLE 331-(3-(1H-imidazol-1-yl)propyl)-3-(4-(methylthio)phenyl)thiourea

melting point: 140.0-140.5° C.

¹H NMR δ 1.8-2.05 (br m, 2H), 2.5 (s, 3H), 3.3-3.5 (br m, 2H), 3.9-4.1(m, 2H), 6.9 (m, 1H), 7.1-7.3 (br m, 5H), 7.6 (s, 1H), 7.75 (br s, 1H),9.4 (s, 1H); MS m/z 307.2 (M+H), 239.2 (M—C₃H₃N₂.)

EXAMPLE 42 1-(3-(1H-imidazol-1-yl)propyl)-3-(4-nitrophenyl)thiourea

melting point: 165.0. 166.0° C.

¹H NMR δ 1.9-2.05 (m, 2H), 3.3-3.5 (br d, 2H), 3.95-4.05 (m, 2H), 6.85(d, 1H), 7.15 (d, 1H), 7.6 (d, 1H), 7.7 (m, 2H), 8.1 (m, 2H), 8.3 (br s,1H), 10.1 (br s, 1H); MS m/z 306.2 (M+H), 237.9 (M—C₃H₃N₂.)

EXAMPLE 501-(3-(1H-imidazol-1-yl)propyl)-3-(4-(dimethylamino)phenyl)thiourea

melting point: 146.5-147.0° C.

¹H NMR δ 1.9-2.0 (m, 2H), 2.9 (s, 6H), 3.4 (m, 2H), 3.9-4.0 (m, 2H), 6.7(m, 2H), 6.9 (s, 1H), 7.05-7.1 (m, 2H), 7.15 (s, 1H), 7.4 (br s, 1H),7.6 (s, 1H), 9.2 (s, 1H); MS m/z 304.2 (M+H), 236.0 (M—C₃H₃N₂.)

EXAMPLE 102 1-(3-(1H-imidazol-1-yl)propyl)-3-(3,4-dimethoxyphenyl)urea

melting point: 114.5-115.0° C.

¹H NMR δ 1.7-1.9 (m, 2H), 2.9-3.1 (m, 2H), 3.7 (2s, 6H), 3.9-4.0 (m,2H), 6.1 (t, 1H), 6.7 (s, 2H), 6.8 (s, 1H), 7.15 (d, 2H), 7.6 (s, 1H),8.2 (s, 1H); MS m/z 321.2 (M+H), 253.3 (M—C₃H₃N₂.)

EXAMPLE 1061-((S)-3-(1H-imidazol-1-yl)-2-methylpropyl)-3-(3,4-dimethoxyphenyl)-thiourea

melting point: 150.5-151.5° C.

¹H NMR δ 0.9 (d, 3H), 2.3-2.4 (m, 2H), 2.5 (s, 1H), 3.7 (d, 6H), 4.0-4.1(br m, 1H), 4.15-4.25 (br m, 1H), 6.75-6.8 (m, 1H), 6.85 (m, 1H),6.9-7.0 (m, 1H), 7.65 (s, 1H), 7.75 (s, 2H), 9.1 (s, 1H), 9.5 (s, 1H);MS m/z 335.6 (M+H), 267.1 (M—C₃H₃N₂.)

EXAMPLE 1071-((R)-3-(1H-imidazol-1-yl)-2-methylpropyl)-3-(3,4-dimethoxyphenyl)-thiourea

melting point: 155.0-157.5° C.

¹H NMR δ 0.9 (d, 3H), 2.3-2.4 (m, 2H), 2.5 (s, 1H), 3.7 (d, 6H), 4.0-4.1(br m, 1H), 4.15-4.25 (br m, 1H), 6.75-6.8 (m, 1H), 6.85 (m, 1H),6.9-7.0 (m, 1H), 7.65 (s, 1H), 7.75 (s, 2H), 9.1 (s, 1H), 9.5 (s, 1H);MS m/z 335.4 (M+H), 267.2 (M—C₃H₃N₂.)

EXAMPLE 1091-((1-((1H-imidazol-1-yl)methyl)cyclopropyl)methyl)-3-(3,4-dimethoxy-phenyl)thiourea

melting point: 166.5-168.5° C.

¹H NMR δ 0.7-0.8 (br m, 2H), 1.85-1.9 (m, 1H), 2.15-2.2 (m, 1H), 2.2-2.3(m, 1H), 3.4-3.5 (m, 1H), 3.7 (d, 6H), 4.2 (s, 1H), 4.95 (s, 1H),6.75-6.8 (br m, 1H), 6.85-6.9 (br m, 1H), 7.0 (s, 1H), 7.5 (m, 1H), 7.6(m, 1H), 7.7 (s, 0.5H), 7.8 (s, 0.5H), 8.85 (s, 0.5H), 9.1 (s, 0.5H),9.35 (s, 0.5H), 9.45 (s, 0.5H); MS m/z 347.2 (M+H), 279.2 (M—C₃H₃N₂.),137.5 (M—C₉H₁₃N₄S.)

EXAMPLE 110 N-(3-(1H-imidazol-1-yl)propyl)benzo[d]thiazol-2-amine

¹H NMR δ 1.95-2.15 (m, 2H), 3.25-3.35 (m, 2H), 4.0-4.1 (t, 2H), 6.9 (s,1H), 6.95-7.05 (t, 1H), 7.15-7.2 (m, 2H), 7.35-7.4 (d, 1H), 7.60-7.70(m, 2H), 8.0-8.1 (br s, 1H); MS m/z 259.4 (M+H), 191.3 (M—C₃H₃N₂.)

EXAMPLE 111N-(3-(1H-imidazol-1-yl)propyl)-6-chlorobenzo[d]thiazol-2-amine

¹H NMR δ 1.95-2.15 (m, 2H), 3.25-3.35 (m, 2H), 4.0-4.1 (t, 2H), 6.9 (s,1H), 7.1-7.2 (d, 2H), 7.3-7.4 (d, 1H), 7.65 (s, 1H), 7.8 (s, 1H), 8.2(s, 1H); MS m/z 293.3 (M+H), 225.3 (M—C₃H₃N₂.)

EXAMPLE 112N-(3-(1H-imidazol-1-yl)propyl)-6-methoxybenzo[d]thiazol-2-amine

¹H NMR δ 1.9-2.05 (m, 2H), 3.2-3.3 (m, 2H), 3.7 (s, 3H), 4.0-4.1 (t,2H), 6.7-6.8 (d, 1H), 6.9 (s, 1H), 7.15-7.2 (s, 1H), 7.2-7.3 (m, 2H),7.65 (s, 1H), 7.8 (s, 1H); MS m/z 289.1 (M+H), 221.4 (M—C₃H₃N₂.)

EXAMPLE 115 (R)—N-(3-(1H-imidazol-1-yl)propyl)-2-phenylpropanethioamide

melting point: 82.0-82.5° C.

¹H NMR δ 1.4-1.55 (d, 3H), 1.9-2.0 (m, 2H), 3.4-3.5 (m, 2H), 3.85-3.95(m, 2H), 4.0-4.1 (q, 1H), 6.8-6.9 (s, 1H), 7.1 (s, 1H), 7.15-7.2 (m,1H), 7.2-7.3 (m, 2H), 7.35-7.4 (m, 2H), 7.55 (s, 1H), 10.1 (s, 1H); MSm/z 274.4 (M+H), 206.3 (M—C₃H₃N₂.)

EXAMPLE 116 (S)—N-(3-(1H-imidazol-1-yl)propyl)-2-phenylpropanethioamide

melting point: 82.5-83.5° C.

¹H NMR δ1.4-1.55 (d, 3H), 1.9-2.0 (m, 2H), 3.4-3.5 (m, 2H), 3.85-3.95(m, 2H), 4.0-4.1 (q, 1H), 6.8-6.9 (s, 1H), 7.1 (s, 1H), 7.15-7.2 (m,1H), 7.2-7.3 (m, 2H), 7.35-7.4 (m, 2H), 7.55 (s, 1H), 10.1 (s, 1H); MSm/z 274.4 (M+H), 206.3 (M—C₃H₃N₂.)

EXAMPLE 121N-(3-(1H-imidazol-1-yl)propyl)-1-(4-chlorophenyl)cyclobutanecarbothioamide

melting point: 137.5-139.0° C.

¹H NMR δ 1.55-1.75 (br m, 2H), 1.85-1.95 (br m, 2H), 2.4-2.5 (br m, 2H),2.7-2.85 (br m, 2H), 3.3-3.5 (br m, 2H), 3.8 (m, 2H), 6.9 (s, 1H), 7.0(s, 1H), 7.3 (m, 2H), 7.45 (s, 1H), 7.5 (m, 2H), 9.6 (t, 1H); MS m/z334.3 (M+H), 266.1 (M—C₃H₃N₂.)

EXAMPLE 122N-(3-(1H-imidazol-1-yl)propyl)-1-(4-chlorophenyl)cyclopentanecarbothioamide

melting point: 140.0-141.0° C.

¹H NMR δ 1.5-1.65 (br m, 4H), 1.8-1.9 (m, 2H), 2.0-2.1 (m, 2H), 2.6 (m,2H), 3.4-3.5 (m, 2H), 3.7-3.8 (m, 2H), 6.85 (s, 1H), 7.0 (s, 1H), 7.35(m, 2H), 7.4 (m, 2H), 7.5 (s, 1H), 9.4 (t, 1H); MS m/z 348.2 (M+H),280.2 (M—C₃H₃N₂.)

EXAMPLE 123N-(3-(1H-imidazol-1-yl)propyl)-1-(4-methoxyphenyl)cyclohexanecarbo-thioamide

melting point: 162.5-164.0° C.

¹H NMR δ 1.2-1.3 (m, 1H), 1.35-1.5 (br m, 5H), 1.85-2.0 (br m, 4H),2.4-2.6 (br m, 2H), 3.4-3.5 (m, 2H), 3.7 (s, 3H), 3.8 (m, 2H), 6.8 (m,3H), 7.0 (s, 1H), 7.3 (m, 2H), 7.5 (s, 1H), 9.2 (t, 1H); MS m/z 358.3(M+H), 290.3 (M—C₃H₃N₂.)

EXAMPLE 124N-(3-(1H-imidazol-1-yl)propyl)-1-(4-methoxyphenyl)cyclopropanecarbothioamide

melting point: 129.0-129.5° C.

¹H NMR δ 1.0-1.1 (m, 2H), 1.5-1.6 (m, 2H), 1.9-2.0 (br m, 2H), 3.4-3.5(m, 2H), 3.7 (s, 3H), 3.9 (m, 2H), 6.9 (m, 3H), 7.1 (s, 1H), 7.2-7.3 (m,2H), 7.6 (s, 1H), 8.9 (br s, 1H); MS m/z 316.0 (M+H), 248.4 (M—C₃H₃N₂.)

EXAMPLE 134 5-(1H-imidazol-1-yl)-N-(3,4-dimethoxyphenyl)pentanethioamide

melting point: 128.0-128.5° C.

¹H NMR δ 1.65-1.70 (m, 2H), 1.75-1.80 (m, 2H), 2.7-2.75 (m, 2H), 3.7 (s,3H), 3.75 (s, 3H), 4.0-4.05 (t, 2H), 6.9-7.0 (m, 2H), 7.2 (s, 1H), 7.3(d, 1H), 7.5 (s, 1H), 7.75 (s, 1H), 11.0 (s, 1H); MS m/z 320.2 (M+H),252.2 (M—C₃H₃N₂.)

EXAMPLE 1361-(2-(1H-imidazol-1-yl)ethyl)-3-(3,4-dimethoxyphenyl)thiourea

melting point: 157.5-159.0° C.

¹H NMR δ 3.7 (2 s, 6H), 3.8 (m, 2H), 4.2 (m, 2H), 6.7 (m, 1H), 6.85 (m,1H), 6.9 (m, 2H), 7.15 (s, 1H), 7.5 (br s, 1H), 7.6 (s, 1H), 9.5 (s,1H); MS m/z 307.2 (M+H), 239.1 (M—C₃H₃N₂.)

EXAMPLE 1371-(4-(1H-imidazol-1-yl)butyl)-3-(3,4-dimethoxyphenyl)thiourea 45

melting point: 114.5-116.0° C.

¹H NMR δ 1.4-1.5 (m, 2H), 1.6-1.7 (m, 2H), 3.4-3.5 (m, 2H), 3.6-3.8 (brs, 6H), 3.9-4.0 (m, 2H), 6.7 (m, 1H), 6.9 (m, 2H), 6.95 (s, 1H), 7.2 (s,1H), 7.6 (br s, 1H), 7.7 (s, 1H), 9.3 (s, 1H); MS m/z 335.3 (M+H), 267.1(M—C₃H₃N₂.)

Physiological substrates of QC (EC) in mammals are, e.g. [Glu³] amyloidβ-protein (3-40/42), [Gln³] amyloid β-protein (3-40/42), Gastrin,Neurotensin, FPP, CCL 2, CCL 7, CCL 8, CCL 16, CCL 18, Fractalkine,Orexin A, [Gln³]-glucagon(3-29) and [Gln⁵]-substance P(5-11). Forfurther details see table 1. The compounds and/or combinations accordingto the present invention and pharmaceutical compositions comprising atleast one inhibitor of QC (EC) are useful for the treatment ofconditions that can be treated by modulation of QC activity.

TABLE 1 Amino acid sequences of physiological active peptides with anN-terminal glutamine residue, which is known to be cyclized to finalpGlu Peptide Amino acid sequence Function Gastrin 17 QGPWL EEEEEAYGWM DFGastrin stimulates the Swiss-Prot: P01350 (amide) stomach mucosa toproduce and secrete hydrochloric acid and the pancreas to secrete itsdigestive enzymes. It also stimulates smooth muscle contraction andincreases blood circulation and water secretion in the stomach andintestine. Neurotensin QLYENKPRRP YIL Neurotensin plays an Swiss-Prot:P30990 endocrine or paracrine role in the regulation of fat metabolism.It causes contraction of smooth muscle. FPP QEP amide A tripeptiderelated to thyrotrophin releasing hormone (TRH), is found in seminalplasma. Recent evidence obtained in vitro and in vivo showed that FPPplays an important role in regulating sperm fertility. TRH QHP amide TRHfunctions as a regulator Swiss-Prot: P20396 of the biosynthesis of TSHin the anterior pituitary gland and as a neurotransmitter/neuromodulator in the central and peripheral nervous systems. GnRHQHWSYGL RP(G) amide Stimulates the secretion of Swiss-Prot: P01148gonadotropins; it stimulates the secretion of both luteinizing andfollicle- stimulating hormones. CCL16 (small QPKVPEW VNTPSTCCLK Showschemotactic activity inducible cytokine YYEKVLPRRL VVGYRKALNC forlymphocytes and A16) HLPAIIFVTK RNREVCTNPN monocytes but not Swiss-Prot:O15467 DDWVQEYIKD PNLPLLPTRN neutrophils. Also shows LSTVKIITAKNGQPQLLNSQ potent myelosuppressive activity, suppresses proliferation ofmyeloid progenitor cells. Recombinant SCYA16 shows chemotactic activityfor monocytes and THP-1 monocytes, but not for resting lymphocytes andneutrophils. Induces a calcium flux in THP-1 cells that weredesensitized by prior expression to RANTES. CCL8 (small QPDSVSIPITCCFNVIN Chemotactic factor that inducible cytokine RKIPIQRLESYTRITNIQCP attracts monocytes, A8) KEAVIFKTKR GKEVCADPKE lymphocytes,basophils and Swiss-Prot: P80075 RWVRDSMKHL DQIFQNLKP eosinophils. Mayplay a role in neoplasia and inflammatory host responses. This proteincan bind heparin. CCL2 (small QPDAINA PVTCCYNFTN Chemotactic factor thatinducible cytokine RKISVQRLAS YRRITSSKCP attracts monocytes and A2)KEAVIFKTIV AKEICADPKQ basophils but not neutrophils Swiss-Prot: P13500KWVQDSMDHL DKQTQTPKT or eosinophils. Augments monocyte anti-tumoractivity. Has been implicated in the pathogenesis of diseasescharacterized by monocytic infiltrates, like psoriasis, rheumatoidarthritis or atherosclerosis. May be involved in the recruitment ofmonocytes into the arterial wall during the disease process ofatherosclerosis. Binds to CCR2 and CCR4. CCL18 (small QVGTNKELCCLVYTSWQIP Chemotactic factor that inducible cytokine QKFIVDYSETSPQCPKPGVI attracts lymphocytes but not A18) LLTKRGRQIC ADPNKKWVQKmonocytes or granulocytes. Swiss-Prot: P55774 YISDLKLNA May be involvedin B cell migration into B cell follicles in lymph nodes. Attracts naiveT lymphocytes toward dendritic cells and activated macrophages in lymphnodes, has chemotactic activity for naive T cells, CD4+ and CD8+ T cellsand thus may play a role in both humoral and cell-mediated immunityresponses. Fractalkine QHHGVT KCNITCSKMT The soluble form is(neurotactin) SKIPVALLIH YQQNQASCGK chemotactic for T cells andSwiss-Prot: P78423 RAIILETRQH RLFCADPKEQ monocytes, but not forWVKDAMQHLD RQAAALTRNG neutrophils. The membrane- GTFEKQIGEV KPRTTPAAGGbound form promotes MDESVVLEPE ATGESSSLEP adhesion of those leukocytesTPSSQEAQRA LGTSPELPTG to endothelial cells. May play VTGSSGTRLPPTPKAQDGGP a role in regulating leukocyte VGTELFRVPP VSTAATWQSS adhesionand migration APHQPGPSLW AEAKTSEAPS processes at the TQDPSTQASTASSPAPEENA endothelium. binds to PSEGQRVWGQ GQSPRPENSL CX3CR1.EREEMGPVPA HTDAFQDWGP GSMAHVSVVP VSSEGTPSRE PVASGSWTPK AEEPIHATMDPQRLGVLITP VPDAQAATRR QAVGLLAFLG LLFCLGVAMF TYQSLQGCPR KMAGEMAEGLRYIPRSCGSN SYVLVPV CCL7 (small QPVGINT STTCCYRFIN Chemotactic factorthat inducible cytokine KKIPKQRLES YRRTTSSHCP attracts monocytes and A7)REAVIFKTKL DKEICADPTQ eosinophils, but not Swiss-Prot: P80098 KWVQDFMKHLDKKTQTPKL neutrophils. Augments monocyte anti-tumor activity. Alsoinduces the release of gelatinase B. This protein can bind heparin.Binds to CCR1, CCR2 and CCR3. Orexin A QPLPDCCRQK TCSCRLYELLNeuropeptide that plays a (Hypocretin-1) HGAGNHAAGI LTL significant rolein the Swiss-Prot O43612 regulation of food intake andsleep-wakefulness, possibly by coordinating the complex behavioral andphysiologic responses of these complementary homeostatic functions. Itplays also a broader role in the homeostatic regulation of energymetabolism, autonomic function, hormonal balance and the regulation ofbody fluids. Orexin-A binds to both OX1R and OX2R with a high affinity.Substance P RPK PQQFFGLM Belongs to the tachykinins. Tachykinins areactive peptides which excite neurons, evoke behavioral responses, arepotent vasodilators and secretagogues, and contract (directly orindirectly) many smooth muscles.

Transepithelial transducing cells, particularly the gastrin (G) cell,co-ordinate gastric acid secretion with the arrival of food in thestomach. Recent work showed that multiple active products are generatedfrom the gastrin precursor, and that there are multiple control pointsin gastrin biosynthesis. Biosynthetic precursors and intermediates(progastrin and Gly-gastrins) are putative growth factors; theirproducts, the amidated gastrins, regulate epithelial cell proliferation,the differentiation of acid-producing parietal cells andhistamine-secreting enterochromaffin-like (ECL) cells, and theexpression of genes associated with histamine synthesis and storage inECL cells, as well as acutely stimulating acid secretion. Gastrin alsostimulates the production of members of the epidermal growth factor(EGF) family, which in turn inhibit parietal cell function but stimulatethe growth of surface epithelial cells. Plasma gastrin concentrationsare elevated in subjects with Helicobacter pylori, who are known to haveincreased risk of duodenal ulcer disease and gastric cancer (Dockray, G.J. 1999 J Physiol 15 315-324).

The peptide hormone gastrin, released from antral G cells, is known tostimulate the synthesis and release of histamine from ECL cells in theoxyntic mucosa via CCK-2 receptors. The mobilized histamine induces acidsecretion by binding to the H(2) receptors located on parietal cells.Recent studies suggest that gastrin, in both its fully amidated and lessprocessed forms (progastrin and glycine-extended gastrin), is also agrowth factor for the gastrointestinal tract. It has been establishedthat the major trophic effect of amidated gastrin is for the oxynticmucosa of stomach, where it causes increased proliferation of gastricstem cells and ECL cells, resulting in increased parietal and ECL cellmass. On the other hand, the major trophic target of the less processedgastrin (e.g. glycine-extended gastrin) appears to be the colonic mucosa(Koh, T. J. and Chen, D. 2000 Regul Pept 9337-44).

Neurotensin (NT) is a neuropeptide implicated in the pathophysiology ofschizophrenia that specifically modulates neurotransmitter systemspreviously demonstrated to be misregulated in this disorder. Clinicalstudies in which cerebrospinal fluid (CSF) NT concentrations have beenmeasured revealed a subset of schizophrenic patients with decreased CSFNT concentrations that are restored by effective antipsychotic drugtreatment. Considerable evidence also exists concordant with theinvolvement of NT systems in the mechanism of action of antipsychoticdrugs. The behavioral and biochemical effects of centrally administeredNT remarkably resemble those of systemically administered antipsychoticdrugs, and antipsychotic drugs increase NT neurotransmission. Thisconcatenation of findings led to the hypothesis that NT functions as anendogenous antipsychotic. Moreover, typical and atypical antipsychoticdrugs differentially alter NT neurotransmission in nigrostriatal andmesolimbic dopamine terminal regions, and these effects are predictiveof side effect liability and efficacy, respectively (Binder, E. B. etal. 2001 Biol Psychiatry 50 856-872).

Fertilization promoting peptide (FPP), a tripeptide related tothyrotrophin releasing hormone (TRH), is found in seminal plasma. Recentevidence obtained in vitro and in vivo showed that FPP plays animportant role in regulating sperm fertility. Specifically, FPPinitially stimulates nonfertilizing (uncapacitated) spermatozoa to“switch on” and become fertile more quickly, but then arrestscapacitation so that spermatozoa do not undergo spontaneous acrosomeloss and therefore do not lose fertilizing potential. These responsesare mimicked, and indeed augmented, by adenosine, known to regulate theadenylyl cyclase (AC)/cAMP signal transduction pathway. Both FPP andadenosine have been shown to stimulate cAMP production in uncapacitatedcells but inhibit it in capacitated cells, with FPP receptors somehowinteracting with adenosine receptors and G proteins to achieveregulation of AC. These events affect the tyrosine phosphorylation stateof various proteins, some being important in the initial “switching on,”others possibly being involved in the acrosome reaction itself.Calcitonin and angiotensin II, also found in seminal plasma, havesimilar effects in vitro on uncapacitated spermatozoa and can augmentresponses to FPP. These molecules have similar effects in vivo,affecting fertility by stimulating and then maintaining fertilizingpotential. Either reductions in the availability of FPP, adenosine,calcitonin, and angiotensin II or defects in their receptors contributeto male infertility (Fraser, L. R. and Adeoya-Osiguwa, S. A. 2001 VitamHorm 63, 1-28).

CCL2, CCL7, CCL8, CCL16, CCL18 and fractalkine play an important role inpathophysiological conditions, such as suppression of proliferation ofmyeloid progenitor cells, neoplasia, inflammatory host responses,cancer, psoriasis, rheumatoid arthritis, atherosclerosis, vasculitis,humoral and cell-mediated immunity responses, leukocyte adhesion andmigration processes at the endothelium, inflammatory bowel disease,restenosis, pulmonary fibrosis, pulmonary hypertention, liver fibrosis,liver cirrhosis, nephrosclerosis, ventricular remodeling, heart failure,arteriopathy after organ transplantations and failure of vein grafts.

Several cytotoxic T lymphocyte peptide-based vaccines against hepatitisB, human immunodeficiency virus and melanoma were recently studied inclinical trials. One interesting melanoma vaccine candidate alone or incombination with other tumor antigens, is the decapeptide ELA. Thispeptide is a Melan-A/MART-1 antigen immunodominant peptide analog, withan N-terminal glutamic acid. It has been reported that the amino groupand gamma-carboxylic group of glutamic acids, as well as the amino groupand gamma-carboxamide group of glutamines, condense easily to formpyroglutamic derivatives. To overcome this stability problem, severalpeptides of pharmaceutical interest have been developed with apyroglutamic acid instead of N-terminal glutamine or glutamic acid,without loss of pharmacological properties. Unfortunately compared withELA, the pyroglutamic acid derivative (PyrELA) and also the N-terminalacetyl-capped derivative (AcELA) failed to elicit cytotoxic T lymphocyte(CTL) activity. Despite the apparent minor modifications introduced inPyrELA and AcELA, these two derivatives probably have lower affinitythan ELA for the specific class I major histocompatibility complex.Consequently, in order to conserve full activity of ELA, the formationof PyrELA must be avoided (Beck A. et al. 2001, J Pept Res57(6):528-38.).

Orexin A is a neuropeptide that plays a significant role in theregulation of food intake and sleep-wakefulness, possibly bycoordinating the complex behavioral and physiologic responses of thesecomplementary homeostatic functions. It plays also a role in thehomeostatic regulation of energy metabolism, autonomic function,hormonal balance and the regulation of body fluids.

By administering a QC (EC)-inhibitor and/or a combination according tothe present invention to a mammal it can be possible to prevent oralleviate or treat conditions selected from Alzheimer's disease, DownSyndrome, ulcer disease and gastric cancer with or w/o Helicobacterpylori infections, neoplasia, inflammatory host responses, cancer,melanoma, malign metastasis, psoriasis, rheumatoid arthritis,atherosclerosis, leukocyte adhesion and migration processes in theendothelium, impaired food intake, sleep-wakefulness, impairedhomeostatic regulation of energy metabolism, impaired autonomicfunction, impaired hormonal balance and impaired regulation of bodyfluids.

Furthermore, by administration of a QC (EC)-inhibitor and/or acombination according to the present invention to a mammal it can bepossible to stimulate the proliferation of myeloid progenitor cells.

In addition, the administration of a QC (EC)-inhibitor and/or acombination according to the present invention can lead to suppressionof male fertility.

In a preferred embodiment, the present invention provides a composition,preferably a pharmaceutical composition comprising at least one QC (EC)inhibitor of formula 1 optionally in combination with at least onecompound selected from the group consisting of PEP-inhibitors, LiCl,inhibitors of dipeptidyl aminopeptidases, preferably inhibitors of DP IVor DP IV-like enzymes, NPY-receptor ligands, NPY agonists,acetylcholinesterase (ACE) inhibitors, PIMT enhancers, inhibitors ofbeta secretases, inhibitors of gamma secretases, inhibitors of neutralendopeptidase, inhibitors of Phosphodiesterase-4 (PDE-4), monoamineoxidase (MAO) inhibitors, TNFalpha inhibitors, amyloid protein oramyloid peptide deposition inhibitors, sigma-1 receptor inhibitors andhistamine H3 antagonists.

Further, the present invention provides pharmaceutical compositions e.g.for parenteral, enteral or oral administration, comprising at least oneQC inhibitor of formula 1 optionally in combination with at least onecompound selected from the group consisting of PEP-inhibitors, LiCl,inhibitors of dipeptidyl aminopeptidases, preferably inhibitors of DP IVor DP IV-like enzymes, NPY-receptor ligands, NPY agonists,acetylcholinesterase (ACE) inhibitors, protein isoaspartatecarboxymethyl transferase (PIMT) enhancers, inhibitors of betasecretases, inhibitors of gamma secretases, inhibitors of neutralendopeptidase, inhibitors of Phosphodiesterase-4 (PDE-4), MAOinhibitors, TNFalpha inhibitors, amyloid protein or amyloid peptidedeposition inhibitors, sigma-1 receptor inhibitors and histamine H3antagonists, optionally in combination with customary carriers and/orexcipients.

These combinations provide a particularly beneficial effect onbehavioral conditions and such combinations are therefore shown to beeffective and useful for the treatment of neuronal disorders, e.g.neuronal diseases selected from the group consisting of Alzheimer'sdisease, Down Syndrome, Parkinson disease, Chorea Huntington, pathogenicpsychotic conditions, schizophrenia, impaired food intake,sleep-wakefulness, impaired homeostatic regulation of energy metabolism,impaired autonomic function, impaired hormonal balance, impairedregulation, body fluids, hypertension, fever, sleep dysregulation,anorexia, anxiety related disorders including depression, seizuresincluding epilepsy, drug withdrawal and alcoholism, neurodegenerativedisorders including cognitive dysfunction and dementia.

Accordingly, the invention provides a method for the treatment ofneuronal disorders, e.g. neuronal diseases selected from the groupconsisting of Alzheimer's disease, Down Syndrome, Parkinson disease,Chorea Huntington, pathogenic psychotic conditions, schizophrenia,impaired food intake, sleep-wakefulness, impaired homeostatic regulationof energy metabolism, impaired autonomic function, impaired hormonalbalance, impaired regulation, body fluids, hypertension, fever, sleepdysregulation, anorexia, anxiety related disorders including depression,seizures including epilepsy, drug withdrawal and alcoholism,neurodegenerative disorders including cognitive dysfunction anddementia, which comprises administering of a thereutically effectiveamount said compositions or combinations to a mammal, preferably ahuman.

Accordingly, the invention provides the use of these compositions orcombinations for the preparation of a medicament for the treatment ofneuronal disorders, e.g. neuronal diseases selected from the groupconsisting of Alzheimer's disease, Down Syndrome, Parkinson disease,Chorea Huntington, pathogenic psychotic conditions, schizophrenia,impaired food intake, sleep-wakefulness, impaired homeostatic regulationof energy metabolism, impaired autonomic function, impaired hormonalbalance, impaired regulation, body fluids, hypertension, fever, sleepdysregulation, anorexia, anxiety related disorders including depression,seizures including epilepsy, drug withdrawal and alcoholism,neurodegenerative disorders including cognitive dysfunction anddementia.

The method comprises either co-administration of at least one QCinhibitor of formula 1 and at least one compound selected from the groupconsisting of PEP-inhibitors, LiCl, inhibitors of dipeptidylaminopeptidases, preferably inhibitors of DP IV or DP IV-like enzymes,NPY-receptor ligands, NPY agonists, ACE inhibitors, PIMT enhancers,inhibitors of beta secretases, inhibitors of gamma secretases,inhibitors of neutral endopeptidase, inhibitors of PDE-4, MAOinhibitors, TNFalpha inhibitors, amyloid protein or amyloid peptidedeposition inhibitors, sigma-1 receptor inhibitors and histamine H3antagonists or the sequential administration thereof.

Co-administration includes administration of a formulation whichincludes at least one QC inhibitor of formula 1 and at least onecompound selected from the group consisting of PEP-inhibitors, LiCl,inhibitors of dipeptidyl aminopeptidases, preferably inhibitors of DP IVor DP IV-like enzymes, NPY-receptor ligands, NPY agonists, ACEinhibitors, PIMT enhancers, inhibitors of beta secretases, inhibitors ofgamma secretases, inhibitors of neutral endopeptidase, inhibitors ofPDE-4, MAO inhibitors, TNFalpha inhibitors, amyloid protein or amyloidpeptide deposition inhibitors, sigma-1 receptor inhibitors and histamineH3 antagonists or the essentially simultaneous administration ofseparate formulations of each agent.

Examples of suitable PIMT enhancers are10-aminoaliphatyl-dibenz[b,f]oxepines of the general formula

described in WO 98/15647 and WO 03/057204, respectively,wherein alk is a divalent aliphatic radical, R is an amino group that isunsubstituted or mono- or di-substituted by monovalent aliphatic and/oraraliphatic radicals or disubstituted by divalent aliphatic radicals,and R₁, R₂, R₃ and R₄ are each, independently of the others, hydrogen,lower alkyl, lower alkoxy, halogen or trifluoromethyl.

Further useful according to the present invention are modulators of PIMTactivity of the general formulae I-IV:

wherein the definition of the substituents R¹-R⁵, (R³)p, (R⁶)p, X, Y andZ is described in WO 2004/039773.

WO 98/15647, WO 03/057204 and WO 2004/039773 are incorporated herein intheir entirety and are part of this invention with regard to thesynthesis and use of the compounds described therein.

Suitable inhibitors of beta and/or gamma secretases and compositionscontaining such inhibitors are described, e.g. in GB 2 385 124, GB 2 389113, US 2002-115616, WO 01/87293, WO 03/057165, WO 2004/052348 and WO2004/062652. These references are incorporated herein in their entiretyand are part of this invention with regard to the synthesis, manufactureand use of the compounds and compositions described therein for theinhibition of beta and/or gamma secretases.

A potent selective and cell permeable gamma secretase inhibitor is(5S)-(t-Butoxycarbonylamino)-6-phenyl-(4R)hydroxy-(2R)benzylhexanoyl)-L-leu-L-phe-amidewith the formula:

A potent beta secretase inhibitor is PNU-33312 of the formula:

Suitable PDE-4 inhibitors are, e.g. shown in the table below:

Company Drug Code Structure Celgene Corp CC-002 Celltech Group plc/MerckFrosst L-826141

Celltech Group plc Sch-351591 (D-4396)

DainipponPharmaceutical CoLtd OS-0217

IBFB Pharma GmbH IBFB-130011 IBFB-150007 IBFB-130020 IBFB-140301 ICOSCorp IC-485

Kings College London VMX-554, VMX-565 Memory MEM-1414 PharmaceuticalsCorp MEM-1018 MEM-1091 MEM-1145 Pfizer Inc CI-1044

Pfizer Inc BHN

Schering AG ZK-117137

SmithKline BeechamPharmaceuticals SB-207499 analogs,GSK

A preferred PDE-4-inhibitor is Rolipram.

A suitable MAO-inhibitor is the compound ladostigil of the formula

Suitable histamine H3 antagonists are, e.g. shown in the table below:

Company Drug Structure AbbottLaboratories A-331440

AbbottLaboratories A-349821

Aventis PharmaAG 3874-H1

Berlin FreeUniversity UCL-2173

BioProjet,Societe Civile deRecherche

BioProjet,Societe Civile deRecherche UCL-1470

Daewoong DWP-302 Pharmaceutical Co Ltd GlaxoSmithKline GSK-189254A IncGSK-207040A Gliatech Inc cipralisant

Gliatech Inc GT-2203

HokkaidoUniversity1S,2S)-2-(2-Aminoethyl)-1-(1H-imidazol-4-yl)cyclopropane

Johnson & JNJ-5207852 Johnson Novo NordiskA/S NNC-0038-0000-1049

Schering-PloughResearchInstitute dual H1/H3antagonists

Schering-PloughResearchInstitute Sch-79687

Suitable inhibitors of prolyl endopeptidase (PEP) are, e.g. chemicalderivatives of proline or small peptides containing terminal prolines.Benzyloxycarbonyl-prolyl-prolinal has been shown to be a specifictransition state inhibitor of the enzyme (Wilk, S. and Orloeski, M., J.Neurochem., 41, 69 (1983), Friedman, et al., Neurochem., 42, 237(1984)). N-terminal substitutions of L-proline or L-prolylpyrrolidine(Atack, et al., Eur. J. of Pharm., 205, 157-163 (1991), JP 03 56,460, EP384,341), as well as variations of N-benzyloxycarbonyl (Z) dipeptidescontaining prolinal at the carboxy terminus have been synthesized asprolyl endopeptidase inhibitors (Nishikata, et al., Chem. Pharm. Bull.34(7), 2931-2936 (1986), Baker, A. et al., Bioorganic & Medicinal Chem.Letts., 1(11), 585-590 (1991)). Thioproline, thiazolidine, andoxopyrrolidine substitutions of the core structure have been reported toinhibit prolyl endopeptidase (Tsuru, et al., J. Biochem., 94, 1179(1988), Tsuru, et al., J. Biochem., 104, 580-586 (1988), Saito et al.,J. Enz. Inhib. 5, 51-75 (1991), Uchida, I., et al. PCT Int. Appl. WO 9012,005, JP 03 56,461, JP 03 56,462). Similarly, various modifications ofthe carboxy terminal proline have been made, including variousfluorinated ketone derivatives (Henning, EP 4,912,127). Generalsyntheses of fluorinated ketone derivatives has been described(Angelastro, M. R., et al., Tetrahedron Letters 33(23), 3265-3268(1992)). Other compounds such as chloromethyl ketone derivatives ofacyl-proline or acylpeptide-proline (Z-Gly-Pro-CH₂Cl) have beendemonstrated to inhibit the enzyme by alkylating the enzyme's activesite (Yoshimoto, T., et al., Biochemistry 16, 2942 (1977)).

EP-A-0 286 928 discloses 2-acylpyrrolidine derivatives useful as propylendopeptidase inhibitors.

Further suitable prolyl endopeptidase inhibitors according to thepresent invention are, e.g. Fmoc-Ala-Pyrr-CN and those listed below:

Z-321 ONO-1603 Zeria Pharmaceutical Co Ono Pharmaceutical Co Ltd Ltd

(4R)-3-(indan-2-ylacetyl)-4- (S)-1-[N-(4-chlorobenzyl)-(1-pyrrolidinyl-carbonyl)-1,3- succinamoyl]pyrrolidin-2- thiazolidincarbaldehyd

JTP-4819 S-17092 Japan Tobacco Inc Servier

(S)-2-{[(S).(hydroxyacatyl)- (2S, 3aS, 7aS)-1{[(R,R)-2-1-pyrrolidinyl]carbonyl}-N- phenylcyclopropyl](phenylmethyl)-1-pyrrolidin- carbonyl}-2-[(thiazolidin-3- carboxamidyl)carbonyl]octahydro-1H- indol

Further suitable prolyl endopeptidase inhibitors according to thepresent invention are disclosed in JP 01042465, JP 03031298, JP04208299, WO 0071144, U.S. Pat. No. 5,847,155; JP 09040693, JP 10077300,JP 05331072, JP 05015314, WO 9515310, WO 9300361, EP 0556482, JP06234693, JP 01068396, EP 0709373, U.S. Pat. No. 5,965,556, U.S. Pat.No. 5,756,763, U.S. Pat. No. 6,121,311, JP 63264454, JP 64000069, JP63162672, EP 0268190, EP 0277588, EP 0275482, U.S. Pat. No. 4,977,180,U.S. Pat. No. 5,091,406, U.S. Pat. No. 4,983,624, U.S. Pat. No.5,112,847, U.S. Pat. No. 5,100,904, U.S. Pat. No. 5,254,550, U.S. Pat.No. 5,262,431, U.S. Pat. No. 5,340,832, U.S. Pat. No. 4,956,380, EP0303434, JP 03056486, JP 01143897, JP 1226880, EP 0280956, U.S. Pat. No.4,857,537, EP 0461677, EP 0345428, JP 02275858, U.S. Pat. No. 5,506,256,JP 06192298, EP 0618193, JP 03255080, EP 0468469, U.S. Pat. No.5,118,811, JP 05025125, WO 9313065, JP 05201970, WO 9412474, EP 0670309,EP 0451547, JP 06339390, U.S. Pat. No. 5,073,549, U.S. Pat. No.4,999,349, EP 0268281, U.S. Pat. No. 4,743,616, EP 0232849, EP 0224272,JP 62114978, JP 62114957, U.S. Pat. No. 4,757,083, U.S. Pat. No.4,810,721, U.S. Pat. No. 5,198,458, U.S. Pat. No. 4,826,870, EP 0201742,EP 0201741, U.S. Pat. No. 4,873,342, EP 0172458, JP 61037764, EP0201743, U.S. Pat. No. 4,772,587, EP 0372484, U.S. Pat. No. 5,028,604,WO 9118877, JP 04009367, JP 04235162, U.S. Pat. No. 5,407,950, WO9501352, JP 01250370, JP 02207070, U.S. Pat. No. 5,221,752, EP 0468339,JP 04211648 and WO 9946272, the teachings of which are hereinincorporated by reference in their entirety, especially concerning theseinhibitors, their definition, uses and their production.

Most preferred is the PEP-inhibitor of the formula:

Other suitable compounds that can be used according to the presentinvention in combination with QC-inhibitors are NPY, a NPY mimetic or aNPY agonist or antagonist or a ligand of the NPY receptors.

Preferred according to the present invention are antagonists of the NPYreceptors. Suitable ligands or antagonists of the NPY receptors are3a,4,5,9b-tetrahydro-1 h-benz[e]indol-2-yl amine-derived compounds asdisclosed in WO 00/68197.

NPY receptor antagonists which may be mentioned include those disclosedin European patent applications EP 0 614 911, EP 0 747 357, EP 0 747 356and EP 0 747 378; international patent applications WO 9417035, WO9719911, WO 9719913, WO 9612489, WO 9719914, WO 9622305, WO 9640660, WO9612490, WO 9709308, WO 9720820, WO 9720821, WO 9720822, WO 9720823, WO9719682, WO 9725041, WO 9734843, WO 9746250, WO 9803492, WO 9803493, WO9803494 and WO 9807420; WO 0030674, U.S. Pat. Nos. 5,552,411, 5,663,192and 5,567,714; 6,114,336, Japanese patent application JP 09157253;international patent applications WO 9400486, WO 9312139, WO 9500161 andWO 9915498; U.S. Pat. No. 5,328,899; German patent application DE 393 9797; European patent applications EP 355 794 and EP 355 793; and Japanesepatent applications JP 06116284 and JP 07267988, the disclosures in allof which documents are hereby incorporated by reference. Preferred NPYantagonists include those compounds that are specifically disclosed inthese patent documents. More preferred compounds include amino acid andnon-peptide-based NPY antagonists. Amino acid and non-peptide-based NPYantagonists which may be mentioned include those disclosed in Europeanpatent applications EP 0 614 911, EP 0 747 357, EP 0 747 356 and EP 0747 378; international patent applications WO 9417035, WO 9719911, WO9719913, WO 9612489, WO 9719914, WO 9622305, WO 9640660, WO 9612490, WO9709308, WO 9720820, WO 9720821, WO 9720822, WO 9720823, WO 9719682, WO9725041, WO 9734843, WO 9746250, WO 9803492, WO 9803493, WO 9803494, WO9807420 and WO 9915498; U.S. Pat. Nos. 5,552,411, 5,663,192 and5,567,714; and Japanese patent application JP 09157253. Preferred aminoacid and non-peptide-based NPY antagonists include those compounds thatare specifically disclosed in these patent documents.

Particularly preferred compounds include amino acid-based NPYantagonists. Amino acid-based compounds which may be mentioned includethose disclosed in international patent applications WO 9417035, WO9719911, WO 9719913, WO 9719914 or, preferably, WO 9915498. Preferredamino acid-based NPY antagonists include those that are specificallydisclosed in these patent documents, for example BIBP3226 and,especially,(R)—N2-(diphenylacetyl)-(R)—N-[1-(4-hydroxy-phenyl)ethyl]arginine amide(Example 4 of international patent application WO 9915498).

For the avoidance of doubt, the examples disclosed in each of the abovementioned publications are specifically incorporated herein by referencein their entirety, as individually disclosed compounds, especiallyconcerning their structure, their definition, uses and their production.

Suitable DP IV-inhibitors are those, disclosed e.g. in U.S. Pat. No.6,380,398, U.S. Pat. No. 6,011,155; U.S. Pat. No. 6,107,317; U.S. Pat.No. 6,110,949; U.S. Pat. No. 6,124,305; U.S. Pat. No. 6,172,081; WO9515309, WO 9961431, WO 9967278, WO 9967279, DE 198 34 591, WO 9740832,DE 196 16 486 C₂, WO 9819998, WO 0007617, WO 9938501, WO 9946272, WO9938501, WO 0168603, WO 0140180, WO 0181337, WO 0181304, WO 0155105, WO0202560 and WO 0214271, WO 0204610, WO 02051836, WO 02068420, WO02076450; WO 02083128, WO 0238541, WO 03000180, WO 03000181, WO03000250, WO 03002530, WO 03002531, WO 03002553, WO 03002593, WO03004496, WO 03004498, WO 03024965, WO 03024942, WO 03035067, WO03037327, WO 03035057, WO 03045977, WO 03055881, WO 0368748, WO 0368757,WO 03057666, WO 03057144, WO 03040174, WO 03033524 and WO 03074500, theteachings of which are herein incorporated by reference in theirentirety, especially concerning these inhibitors, their definition, usesand their production.

Further suitable DP IV-inhibitors are, e.g. shown in the table below:

Company Drug Code Structure Bristol-Myers Squibb Co BMS-477118

Eisai Co Ltd

Ferring Research Ltd FE-999011

GlaxoSmithKline plc GW-229A

Kyowa Hakko Kogyo K-579

Merck & Co Inc MK-431

Novartis AG LAF-237

Novo Nordisk A/S

Novo Nordisk A/S Valine pyrrolidine Pfizer Inc CP-867534-01

Phenomix Corp. PHX-1004 Point Therapeutics Inc. PT-100(Talabostat)

Sanofi-Synthelabo SSR-162369 Syrrx Inc SYR-322 Taisho PharmaceuticalCoLtd

Tanabe Seiyaku Co Ltd TSL-225

Tanabe Seiyaku Co 815541 Ltd., licensed to GlaxoSmithKline plc

For the avoidance of doubt, the references and examples disclosed hereinare specifically incorporated herein by reference in their entirety, oras individually disclosed compounds, especially concerning theirstructure, their definition, uses and their production.

Preferred DP IV-inhibitors are dipeptide-like compounds and compoundsanalogous to dipeptide compounds that are formed from an amino acid anda thiazolidine or pyrrolidine group, and salts thereof, referred tohereinafter as dipeptide-like compounds. Preferably the amino acid andthe thiazolidine or pyrrolidine group are bonded with an amide bond.

Especially suitable for that purpose according to the invention aredipeptide-like compounds in which the amino acid is preferably selectedfrom a natural amino acid, such as, for example, leucine, valine,glutamine, glutamic acid, proline, isoleucine, asparagines and asparticacid.

The dipeptide-like compounds used according to the invention exhibit ata concentration (of dipeptide compounds) of 10 μM, a reduction in theactivity of plasma dipeptidyl peptidase IV or DP IV-analogous enzymeactivities of at least 10%, especially of at least 40%. Frequently areduction in activity of at least 60% or at least 70% is also required.Preferred agents may also exhibit a reduction in activity of a maximumof 20% or 30%.

Preferred dipeptide-like compounds are N-valyl prolyl, O-benzoylhydroxylamine, alanyl pyrrolidine, isoleucyl thiazolidine likeL-allo-isoleucyl thiazolidine, L-threo-isoleucyl pyrrolidine and saltsthereof, especially the fumaric salts, and L-allo-isoleucyl pyrrolidineand salts thereof.

Further preferred compounds are given in Table 2.

The salts of the dipeptide-like compounds can be present in a molarratio of dipeptide (-analogous) component to salt component of 1:1 or2:1. Such a salt is, for example, (Ile-Thia)₂ fumaric acid.

TABLE 2 Structures of further preferred dipeptide compounds DPIV-inhibitor H-Asn-pyrrolidine H-Asn-thiazolidine H-Asp-pyrrolidineH-Asp-thiazolidine H-Asp(NHOH)-pyrrolidine H-Asp(NHOH)-thiazolidineH-Glu-pyrrolidine H-Glu-thiazolidine H-Glu(NHOH)-pyrrolidineH-Glu(NHOH)-thiazolidine H-His-pyrrolidine H-His-thiazolidineH-Pro-pyrrolidine H-Pro-thiazolidine H-Ile-azididine H-Ile-pyrrolidineH-L-allo-Ile-thiazolidine H-Val-pyrrolidine H-Val-thiazolidine

In another preferred embodiment, the present invention provides the useof compounds of formula 3 for competitive modulation of dipeptidylpeptidase IV catalysis:

whereinA, B, C, D and E are independently any amino acid moieties includingproteinogenic amino acids, non-proteinogenic amino acids, L-amino acidsand D-amino acids and wherein E and/or D may be absent.

According to preferred embodiments, the residues A, B, C, D and E offormula (3) are independently defined as follows:

-   -   A is an amino acid except a D-amino acid,    -   B is an amino acid selected from Pro, Ala, Ser, Gly, Hyp,        acetidine-(2)-carboxylic acid and pipecolic acid,    -   C is any amino acid except Pro, Hyp, acetidine-(2)-carboxylic        acid, pipecolic acid and except N-alkylated amino acids, e.g.        N-methyl valine and sarcosine,    -   D is any amino acid or missing, and    -   E is any amino acid or missing,        or:    -   C is any amino acid except Pro, Hyp, acetidine-(2)-carboxylic        acid, pipecolic acid, except N-alkylated amino acids, e.g.        N-methyl valine and sarcosine, and except a D-amino-acid;    -   D is any amino acid selected from Pro, Ala, Ser, Gly, Hyp,        acetidine-(2)-carboxylic acid and pipecolic acid, and    -   E is any amino acid except Pro, Hyp, acetidine-(2)-carboxylic        acid, pipecolic acid and except N-alkylated amino acids, e.g.        N-methyl valine and sarcosine.

Other amino acids than those encoded in the genetic code can also beincluded in peptide compounds within the scope of the invention and canbe classified within this general scheme.

Proteinogenic amino acids are defined herein as natural protein-derivedα-amino acids. Non-proteinogenic amino acids are defined herein as allother amino acids, which are not building blocks of common naturalproteins.

The resulting peptides may be synthesized as the free C-terminal acid oras the C-terminal amide form. The free acid peptides or the amides maybe varied by side chain modifications. Such side chain modificationsinclude for instance, but are not restricted to, homoserine formation,pyroglutamic acid formation, disulphide bond formation, deamidation ofasparagine or glutamine residues, methylation, t-butylation,t-butyloxycarbonylation, 4-methylbenzylation, thioanysilation,thiocresylation, benzyloxymethylation, 4-nitrophenylation,benzyloxycarbonylation, 2-nitrobencoylation, 2-nitrosulphenylation,4-toluenesulphonylation, pentafluorophenylation, diphenylmethylation,2-chlorobenzyloxycarbonylation, 2,4,5-trichlorophenylation,2-bromobenzyloxycarbonylation, 9-fluorenylmethyloxycarbonylation,triphenylmethylation, 2,2,5,7,8,-pentamethylchroman-6-sulphonylation,hydroxylation, oxidation of methionine, formylation, acetylation,anisylation, benzylation, bencoylation, trifluoroacetylation,carboxylation of aspartic acid or glutamic acid, phosphorylation,sulphation, cysteinylation, glycolysation with pentoses, deoxyhexoses,hexosamines, hexoses or N-acetylhexosamines, farnesylation,myristolysation, biotinylation, palmitoylation, stearoylation,geranylgeranylation, glutathionylation, 5′-adenosylation,ADP-ribosylation, modification with N-glycolylneuraminic acid,N-acetylneuraminic acid, pyridoxal phosphate, lipoic acid,4′-phosphopantetheine, or N-hydroxysuccinimide.

In the compounds of formula (3), the amino acid moieties A, B, C, D, andE are respectively attached to the adjacent moiety by amide bonds in ausual manner according to standard nomenclature so that theamino-terminus (N-terminus) of the amino acids (peptide) is drawn on theleft and the carboxyl-terminus of the amino acids (peptide) is drawn onthe right. (C-terminus).

Preferred peptide compounds are listed in table 3.

TABLE 3 Examples of peptide substrates Mass (exp.)¹ Peptide Mass (calc.)[M + H⁺] 2-Amino octanoic acid-Pro-Ile 369.5 370.2 Abu-Pro-Ile 313.4314.0 Aib-Pro-Ile 313.4 314.0 Aze-Pro-Ile 311.4 312.4 Cha-Pro-Ile 381.52382.0 Ile-Hyp-Ile 356.45 358.2 Ile-Pro-allo-Ile 341.4 342.0Ile-Pro-t-butyl-Gly 341.47 342.36 Ile-Pro-Val 327.43 328.5 Nle-Pro-Ile341.45 342.2 Nva-Pro-Ile 327.43 328.2 Orn-Pro-Ile 342.42 343.1Phe-Pro-Ile 375.47 376.2 Phg-Pro-Ile 361.44 362.2 Pip-Pro-Ile 338.56340.0 Ser(Bzl)-Pro-Ile 405.49 406.0 Ser(P)-Pro-Ile 395.37 396.0Ser-Pro-Ile 315.37 316.3 t-butyl-Gly-Pro-D-Val 327.4 328.6t-butyl-Gly-Pro-Gly 285.4 286.3 t-butyl-Gly-Pro-Ile 341.47 342.1t-butyl-Gly-Pro-Ile-amide 340.47 341.3 t-butyl-Gly-Pro-t-butyl-Gly341.24 342.5 t-butyl-Gly-Pro-Val 327.4 328.4 Thr-Pro-Ile 329.4 330.0Tic-Pro-Ile 387.46 388.0 Trp-Pro-Ile 414.51 415.2 Tyr(P)-Pro-Ile 471.47472.3 Tyr-Pro-allo-Ile 391.5 392.0 Val-Pro-allo-Ile 327.4 328.5Val-Pro-t-butyl-Gly 327.4 328.15 Val-Pro-Val 313.4 314.0¹[M+H⁺] were determined by Electrospray mass spectrometry in positiveionization mode.

t-butyl-Gly is defined as:

Ser(Bzl) and Ser(P) are defined as benzyl-serine and phosphoryl-serine,respectively. Tyr(P) is defined as phosphoryl-tyrosine.

Further preferred DP IV-inhibitors, which can be used according to thepresent invention, are peptidylketones of formula 4:

and pharmaceutically acceptable salts thereof, wherein:A is selected from the following structures:

-   -   wherein    -   X¹ is H or an acyl or oxycarbonyl group including an amino acid        residue, N-protected amino acid residue, a peptide residue or a        N-protected peptide residue,    -   X² is H, —(CH)_(m)—NH—C₅H₃N—Y with m=2-4 or —C₅H₃N—Y (a divalent        pyridyl residue) and Y is selected from H, Br, Cl, I, NO₂ or CN,    -   X³ is H or selected from an alkyl-, alkoxy-, halogen-, nitro-,        cyano- or carboxy-substituted phenyl or from an alkyl-, alkoxy-,        halogen-, nitro-, cyano- or carboxy-substituted pyridyl residue,    -   X⁴ is H or selected from an alkyl-, alkoxy-, halogen-, nitro-,        cyano- or carboxy-substituted phenyl or from an alkyl-, alkoxy-,        halogen-, nitro-, cyano- or carboxy-substituted pyridyl residue,    -   X⁵ is H or an alkyl, alkoxy or phenyl residue,    -   X⁶ is H or an alkyl residue,    -   for n=1        X is selected from: H, OR², SR², NR²R³, N⁺R²R³R⁴, wherein:    -   R² stands for acyl residues, which are optionally substituted        with alkyl, cycloalkyl, aryl or heteroaryl residues, or for        amino acid residues or peptidic residues, or alkyl residues,        which are optionally substituted with alkyl, cycloalkyl, aryl or        heteroaryl residues,    -   R³ stands for alkyl or acyl residues, wherein R² and R³ may be        part of a saturated or unsaturated carbocyclic or heterocyclic        ring,    -   R⁴ stands for alkyl residues, wherein R² and R⁴ or R³ and R⁴ may        be part of a saturated or unsaturated carbocyclic or        heterocyclic ring,    -   for n=0        X is selected from:

-   -   wherein    -   B stands for: O, S or NR⁵, wherein R⁵ is H, alkyl or acyl,    -   C, D, E, F, G, Y, K, L, M, Q, T, U, V and W are independently        selected from alkyl and substituted alkyl residues, oxyalkyl,        thioalkyl, aminoalkyl, carbonylalkyl, acyl, carbamoyl, aryl and        heteroaryl residues, and        Z is selected from H, or a branched or straight chain alkyl        residue from C₁-C₉, a branched or straight chain alkenyl residue        from C₂-C₉, a cycloalkyl residue from C₃-C₈, a cycloalkenyl        residue from C₅-C₇, an aryl or heteroaryl residue, or a side        chain selected from all side chains of all natural amino acids        or derivatives thereof.

In preferred compounds of formula 4, A is

-   -   wherein    -   X¹ is H or an acyl or oxycarbonyl group including an amino acid        residue, N-acylated amino acid residue, a peptide residue from        di- to pentapeptides, preferably a dipeptide residue, or a        N-protected peptide residue from di- to pentapeptides,        preferably a N-protected dipeptide residue    -   X² is H, —(CH)_(m)—NH—C₅H₃N—Y with m=2-4 or —C₅H₃N—Y (a divalent        pyridyl residue) and Y is selected from H, Br, Cl, I, NO₂ or CN,    -   for n=1        X is preferably selected from: H, OR², SR², NR²R³, wherein:    -   R² stands for acyl residues, which are optionally substituted        with alkyl, cycloalkyl, aryl or heteroaryl residues, or for        amino acid residues or peptidic residues, or alkyl residues,        which are optionally substituted with alkyl, cycloalkyl, aryl or        heteroaryl residues,    -   R³ stands for alkyl or acyl residues, wherein R² and R³ may be        part of a saturated or unsaturated carbocyclic or heterocyclic        ring,    -   for n=0        X is preferably selected from:

-   -   wherein    -   B stands for: O, S or NR⁵, wherein R⁵ is H, alkyl or acyl,    -   C, D, E, F, G, Y, K, L, M and Q are independently selected from        alkyl and substituted alkyl residues, oxyalkyl, thioalkyl,        aminoalkyl, carbonylalkyl, acyl, carbamoyl, aryl and heteroaryl        residues, and        Z is selected from H, or a branched or straight chain alkyl        residue from C₁-C₉, preferably C₂-C₆, a branched or straight        chain alkenyl residue from C₂-C₉, a cycloalkyl residue from        C₃-C₈, a cycloalkenyl residue from C₅-C₇, an aryl or heteroaryl        residue, or a side chain selected from all side chains of all        natural amino acids or derivatives thereof.

In more preferred compounds of formula 4, A is

-   -   wherein    -   X¹ is H or an acyl or oxycarbonyl group including an amino acid        residue, N-acylated amino acid residue or a peptide residue from        di- to pentapeptides, preferably a dipeptide residue, or a        N-protected peptide residue from di- to pentapeptides,        preferably a N-protected dipeptide residue    -   for n=1,        X is preferably selected from: H, OR², SR², wherein:    -   R² stands for acyl residues, which are optionally substituted        with alkyl or aryl residues,    -   for n=0        X is preferably selected from:

-   -   wherein    -   B stands for: O, S or NR⁵, wherein R⁵ is H, alkyl or acyl,    -   C, D, E, F, G, Y, K, L, M and Q are independently selected from        alkyl and substituted alkyl residues, oxyalkyl, thioalkyl,        aminoalkyl, carbonylalkyl, acyl, carbamoyl, aryl and heteroaryl        residues, and        Z is selected from H, or a branched or straight chain alkyl        residue from C₁-C₉, preferably C₂-C₆, a branched or straight        chain alkenyl residue from C₂-C₉, a cycloalkyl residue from        C₃-C₈, a cycloalkenyl residue from C₅-C₇, an aryl or heteroaryl        residue, or a side chain selected from all side chains of all        natural amino acids or derivatives thereof.

In most preferred compounds of formula 4, A is

-   -   wherein    -   X¹ is H or an acyl or oxycarbonyl group including an amino acid        residue, N-acylated amino acid residue or a dipeptide residue,        containing a Pro or Ala in the penultimate position, or a        N-protected dipeptide residue containing a Pro or Ala in the        penultimate position,    -   for n=1,

X is H,

-   -   for n=0        X is preferably selected from:

-   -   wherein    -   B stands for: O or S, most preferably for S    -   C, D, E, F, G, Y, K, L, M, Q, are H and        Z is selected from H, or a branched or straight chain alkyl        residue from C₃-C₅, a branched or straight chain alkenyl residue        from C₂-C₉, a cycloalkyl residue from C₅-C₇, a cycloalkenyl        residue from C₅-C₇, an aryl or heteroaryl residue, or a side        chain selected from all side chains of all natural amino acids        or derivatives thereof.

Most preferred for Z is H.

According to a preferred embodiment the acyl groups are C₁-C₆-acylgroups.

According to a further preferred embodiment the alk(yl) groups areC₁-C₆-alk(yl) groups, which may be branched or unbranched.

According to a still further preferred embodiment the alkoxy groups areC₁-C₆-alkoxy groups.

According to yet another preferred embodiment the aryl residues areC₅-C₁₂ aryl residues that have optionally fused rings.

According to a still further preferred embodiment the cycloalkylresidues (carbocycles) are C₃-C₈-cycloalkyl residues.

According to another preferred embodiment the heteroaryl residues areC₄-C₁₁ aryl residues that have optionally fused rings and, in at leastone ring, additionally from 1 to 4 preferably 1 or 2 hetero atoms, suchas O, N and/or S.

According to a further preferred embodiment peptide residues containfrom 2 to 50 amino acids.

According to another preferred embodiment the heterocyclic residues areC₂-C₇-cycloalkyl radicals that additionally have from 1 to 4, preferably1 or 2 hetero atoms, such as O, N and/or S.

According to a still further preferred embodiment the carboxy groups areC₁-C₆ carboxy groups, which may be branched or unbranched.

According to yet another preferred embodiment the oxycarbonyl groups aregroups of the formula —O—(CH₂)₁₋₆COOH.

The amino acids can be any natural or synthetic amino acid, preferablynatural alpha amino acids.

Preferred compounds of formula (4) are2-Methylcarbonyl-1-N-[(L)-Alanyl-(L)-Valinyl]-(2S)-pyrrolidinehydrobromide;2-Methyl)carbonyl-1-N-[(L)-Valinyl-(L)-Prolyl-(L)-Valinyl]-(2S)-pyrrolidinehydrobromide;2-[(Acetyl-oxy-methyl)carbonyl]-1-N-[(L)-Alanyl-(L)-Valinyl]-(2S)-pyrrolidinehydrobromide;2-[Benzoyl-oxy-methyl)carbonyl]-1-N-[{(L)-Alanyl}-(L)-Valinyl]-(2S)-pyrrolidinehydrobromide;2-{[(2,6-Dichlorbenzyl)thiomethyl]carbonyl}-1-N-[{(L)-Alanyl}-(L)-Valinyl]-(2S)-pyrrolidine;2-[Benzoy-loxy-methyl)carbonyl]-1-N-[Glycyl-(L)-Valinyl]-(2S)-pyrrolidinehydrobromide;2-[([1,3]-thiazole-2-yl)carbonyl]-1-N-[{(L)-Alanyl}-(L)-Valinyl]-(2S)-pyrrolidinetrifluoracetat;2-[(benzothiazole-2-yl)carbonyl]-1-N—[N-{(L)-Alanyl}-(L)-Valinyl]-(2S)-pyrrolidintrifluoracetat;2-[(-benzothiazole-2-yl)carbonyl]-1-N-[{(L)-Alanyl}-Glycyl]-(2S)-pyrrolidinetrifluoracetat;2-[(pyridin-2-yl)carbonyl]-1-N—[N-{(L)-Alanyl}-(L)-Valinyl]-(2S)-pyrrolidinetrifluoracetat.

Further, according to the present invention preferred DP IV-inhibitorsare compounds of formula (5) including all stereoisomers andpharmaceutical acceptable salts:

B—(CH—R¹)_(n)—C(═X²)-D  (5)

whereinn is 0 or 1,R¹ stands for H, C₁-C₉ branched or straight chain alkyl, preferably H,n-butan-2-yl, n-prop-2-yl or isobutyl, C₂-C₉ branched or straight chainalkenyl, C₃-C₈ cycloalkyl, preferably cyclohexyl, C₅-C₇ cycloalkenyl,aryl, heteroaryl or a side chain of a natural amino acid or mimeticsthereof,X² stands for O, NR⁶, N⁺(R⁷)₂, or S,B is selected from the following groups:

-   -   where X⁵ is H or an acyl or oxycarbonyl group including amino        acids,    -   R⁵ is H, C₁-C₉ branched or straight chain alkyl, preferably H,        n-butan-2-yl, n-prop-2-yl or isobutyl, C₂-C₉ branched or        straight chain alkenyl, C₃-C₈ cycloalkyl, preferably cyclohexyl,        3-hydroxyadamant-1-yl, C₅-C₇ cycloalkenyl, aryl, heteroaryl or a        side chain of a natural amino acid or derivatives thereof, or a        group of the formula —(CH)_(m)—NH—C₅H₃N—Y where m is an integer        of 2-4, —C₅H₃N—Y is a divalent pyridyl moiety and Y is a        hydrogen atom, a halogen atom, a nitro group or a cyano group,    -   R⁶, R⁷ and R⁸ are independently selected from H, optionally        substituted C₁-C₉ branched or straight chain alkyl, preferably        an optionally substituted C₂-C₅ branched or straight chain        alkyl; or optionally substituted C₂-C₉ branched or straight        chain alkenyl, preferably an C₂-C₅ branched or straight chain        alkenyl; or optionally substituted C₃-C₈ cycloalkyl, preferably        an optionally substituted C₄-C₇ cycloalkyl; or an optionally        substituted C₅-C₇ cycloalkenyl, or an optionally substituted        aryl residue,    -   Z is selected from H, pyridyl or optionally substituted phenyl,        optionally substituted alkyl groups, alkoxy groups, halogens,        nitro, cyano and carboxy groups,    -   W is selected from H, pyridyl or optionally substituted phenyl,        optionally substituted alkyl groups, alkoxy groups, halogens,        nitro, cyano and carboxy groups,    -   W¹ is H or optionally substituted alkyl, alkoxy or optionally        substituted phenyl, and    -   Z¹ is H, or optionally substituted alkyl,    -   R³ and R⁴ are independently H, hydroxy, alkyl, alkoxy, aralkoxy,        nitro, cyano or halogen,        D is an optionally substituted compound of the formula

which can be saturated, or can have one, two or three double bonds,wherein

-   -   X⁸ to X¹¹ are independently CH, N, N⁺(R⁷), or CR⁸, if        unsaturated, or    -   X⁸ to X¹¹ are independently CH₂, NH, NH⁺(R⁷), O, or S if        saturated,    -   X¹² is CHA, NA, CH₂, NH, NH⁺(R⁷), or CHR⁸, if saturated or    -   X¹² is CA, NA⁺, CH, N, N⁺(R⁷), or CR⁸, if unsaturated and    -   A is H or an isoster of a carboxylic acid such as CN, SO₃H,        CONOH, PO₃R⁵R⁶, a tetrazole, an amide, an ester or an acid        anhydride.

Throughout the application, D contains preferably at most two, furtherpreferred at most one hetero atom in the ring.

According to preferred embodiments of the present invention, D standsfor optionally substituted C₄-C₇ cycloalkyl, preferably C₄-C₆cycloalkyl, optionally substituted C₄-C₇ cycloalkenyl, or optionallysubstituted (hetero)cycloalkyl of the formulae

wherein the residues are as defined above,or

that is, a five-membered ring containing one or two double bonds in thering,wherein the residues are as defined above,or

wherein the residues are as defined above,or

wherein the residues are as defined above,or

that is a six-membered ring containing one or two double bonds in thering,wherein the residues are as defined above,or

wherein the residues are as defined above.

According to a preferred embodiment, B has the following formula:

wherein the residues are as defined above.

According to another preferred embodiment, B has the following formula:

wherein the residues are as defined above.

Preferred compounds according to formula (5) are

-   1-cyclopentyl-3-methyl-1-oxo-2-pentanaminium chloride,-   1-cyclopentyl-3-methyl-1-oxo-2-butanaminium chloride,-   1-cyclopentyl-3,3-dimethyl-1-oxo-2-butanaminium chloride,-   1-cyclohexyl-3,3-dimethyl-1-oxo-2-butanaminium chloride,-   3-(cyclopentylcarbonyl)-1,2,3,4-tetrahydroisoquinolinium chloride,    and-   N-(2-cyclopentyl-2-oxoethyl)cyclohexanaminium chloride.

Because of the wide distribution of the protein in the body and the widevariety of mechanisms involving DP IV, DP IV-activity and DP IV-relatedproteins, systemic therapy (enteral or parenteral administration) withDP IV-inhibitors can result in a series of undesirable side-effects.

The problem to be solved was therefore moreover, to provide DPIV-inhibitors that can be used in combination therapy of neuronaldiseases, for targeted influencing of locally limitedpatho-physiological and physiological processes. The problem of theinvention especially consists in obtaining locally limited and highlyspecific inhibition of DP IV or DP IV-analogous activity for the purposeof targeted intervention in the regulation of the activity of locallyactive substrates.

This problem is solved according to the invention by the use of the DPIV-inhibitors of the general formula (6):

whereinA is an amino acid having at least one functional group in the sidechain,B is a chemical compound covalently bound to at least one functionalgroup of the side chain of A,C is a thiazolidine, pyrrolidine, cyanopyrrolidine, hydroxyproline,dehydroproline or piperidine group amide-bonded to A.

In accordance with a preferred embodiment of the invention,pharmaceutical compositions are used comprising at least one compound ofthe general formula (5) and at least one customary adjuvant appropriatefor the site of action.

Preferably A is an α-amino acid, especially a natural α-amino acidhaving one, two or more functional groups in the side chain, preferablythreonine, tyrosine, serine, arginine, lysine, aspartic acid, glutamicacid or cysteine.

Preferably B is an oligopeptide having a chain length of up to 20 aminoacids, a polyethylene glycol having a molar mass of up to 20 000 g/mol,an optionally substituted organic amine, amide, alcohol, acid oraromatic compound having from 8 to 50 C atoms.

Despite an extended side chain function, the compounds of formula (6)can still bind to the active centre of the enzyme dipeptidyl peptidaseIV and analogous enzymes but are no longer actively transported by thepeptide transporter PepT1. The resulting reduced or greatly restrictedtransportability of the compounds according to the invention leads tolocal or site directed inhibition of DP IV and DP IV-like enzymeactivity.

By extending/expanding the side chain modifications, for example beyonda number of seven carbon atoms, it is accordingly possible to obtain adramatic reduction in transportability. With increasing spatial size ofthe side chains, there is a reduction in the transportability of thesubstances. By spatially and sterically expanding the side chains, forexample beyond the atom group size of a monosubstituted phenyl radical,hydroxylamine radical or amino acid residue, it is possible according tothe invention to modify or suppress the transportability of the targetsubstances.

Preferred compounds of formula (6) are compounds, wherein theoligopeptides have chain lengths of from 3 to 15, especially from 4 to10, amino acids, and/or the polyethylene glycols have molar masses of atleast 250 g/mol, preferably of at least 1500 g/mol and up to 15 000g/mol, and/or the optionally substituted organic amines, amides,alcohols, acids or aromatic compounds have at least 12 C atoms andpreferably up to 30 C atoms.

Pharmaceutical Compositions

To prepare the pharmaceutical compositions of this invention, at leastone effector of QC optionally in combination with at least onePEP-inhibitor and/or at least one DP IV-inhibitor and/or at least oneNPY-receptor-ligand and/or at least one ACE-inhibitor, can be used asthe active ingredient(s). The active ingredient(s) is intimately admixedwith a pharmaceutical carrier according to conventional pharmaceuticalcompounding techniques, which carrier may take a wide variety of formsdepending of the form of preparation desired for administration, e.g.,oral or parenteral such as intramuscular. In preparing the compositionsin oral dosage form, any of the usual pharmaceutical media may beemployed. Thus, for liquid oral preparations, such as for example,suspensions, elixirs and solutions, suitable carriers and additivesinclude water, glycols, oils, alcohols, flavoring agents, preservatives,coloring agents and the like; for solid oral preparations such as, forexample, powders, capsules, gelcaps and tablets, suitable carriers andadditives include starches, sugars, diluents, granulating agents,lubricants, binders, disintegrating agents and the like. Because oftheir ease in administration, tablets and capsules represent the mostadvantageous oral dosage unit form, in which case solid pharmaceuticalcarriers are obviously employed. If desired, tablets may be sugar coatedor enteric coated by standard techniques. For parenterals, the carrierwill usually comprise sterile water, though other ingredients, forexample, for purposes such as aiding solubility or for preservation, maybe included.

Injectable suspensions may also prepared, in which case appropriateliquid carriers, suspending agents and the like may be employed. Thepharmaceutical compositions herein will contain, per dosage unit, e.g.,tablet, capsule, powder, injection, teaspoonful and the like, an amountof the active ingredient(s) necessary to deliver an effective dose asdescribed above. The pharmaceutical compositions herein will contain,per dosage unit, e.g., tablet, capsule, powder, injection, suppository,teaspoonful and the like, from about 0.03 mg to 100 mg/kg (preferred0.1-30 mg/kg) and may be given at a dosage of from about 0.1-300 mg/kgper day (preferred 1-50 mg/kg per day) of each active ingredient orcombination thereof. The dosages, however, may be varied depending uponthe requirement of the patients, the severity of the condition beingtreated and the compound being employed. The use of either dailyadministration or post-periodic dosing may be employed.

Preferably these compositions are in unit dosage forms from such astablets, pills, capsules, powders, granules, sterile parenteralsolutions or suspensions, metered aerosol or liquid sprays, drops,ampoules, autoinjector devices or suppositories; for oral parenteral,intranasal, sublingual or rectal administration, or for administrationby inhalation or insufflation. Alternatively, the composition may bepresented in a form suitable for once-weekly or once-monthlyadministration; for example, an insoluble salt of the active compound,such as the decanoate salt, may be adapted to provide a depotpreparation for intramuscular injection. For preparing solidcompositions such as tablets, the principal active ingredient is mixedwith a pharmaceutical carrier, e.g. conventional tableting ingredientssuch as corn starch, lactose, sucrose, sorbitol, talc, stearic acid,magnesium stearate, dicalcium phosphate or gums, and otherpharmaceutical diluents, e.g. water, to form a solid preformulationcomposition containing a homogeneous mixture of a compound of thepresent invention, or a pharmaceutically acceptable salt thereof. Whenreferring to these preformulation compositions as homogeneous, it ismeant that the active ingredient is dispersed evenly throughout thecomposition so that the composition may be readily subdivided intoequally effective dosage forms such as tablets, pills and capsules. Thissolid preformulation composition is then subdivided into unit dosageforms of the type described above containing from 0.1 to about 500 mg ofeach active ingredient or combinations thereof of the present invention.

The tablets or pills of the compositions of the present invention can becoated or otherwise compounded to provide a dosage form affording theadvantage of prolonged action. For example, the tablet or pill cancomprise an inner dosage and an outer dosage component, the latter beingin the form of an envelope over the former. The two components can beseparated by an enteric layer which serves to resist disintegration inthe stomach and permits the inner component to pass intact into theduodenum or to be delayed in release. A variety of material can be usedfor such enteric layers or coatings, such materials including a numberof polymeric acids with such materials as shellac, cetyl alcohol andcellulose acetate.

This liquid forms in which the compositions of the present invention maybe incorporated for administration orally or by injection include,aqueous solutions, suitably flavoured syrups, aqueous or oilsuspensions, and flavoured emulsions with edible oils such as cottonseedoil, sesame oil, coconut oil or peanut oil, as well as elixirs andsimilar pharmaceutical vehicles. Suitable dispersing or suspendingagents for aqueous suspensions, include synthetic and natural gums suchas tragacanth, acacia, alginate, dextran, sodium carboxymethylcellulose,methylcellulose, polyvinylpyrrolidone or gelatin.

Where the processes for the preparation of the compounds of the presentinvention give rise to mixture of stereoisomers, these isomers may beseparated by conventional techniques such as preparative chromatography.The compounds may be prepared in racemic form, or individual enantiomersmay be prepared either by enantiospecific synthesis or by resolution.The compounds may, for example, be resolved into their componentsenantiomers by standard techniques, such as the formation ofdiastereomeric pairs by salt formation with an optically active acid,such as (−)-di-p-toluoyl-d-tartaric acid and/or(+)-di-p-toluoyl-l-tartaric acid followed by fractional crystallizationand regeneration of the free base. The compounds may also resolved byformation of diastereomeric esters or amides, followed bychromatographic separation and removal of the chiral auxiliary.Alternatively, the compounds may be resolved using a chiral HPLC column.

During any of the processes for preparation of the compounds of thepresent invention, it may be necessary and/or desirable to protectsensitive or reactive groups on any of the molecules concerned. This maybe achieved by means of conventional protecting groups, such as thosedescribed in Protective Groups in Organic Chemistry, ed. J. F. W.McOmie, Plenum Press, 1973; and T. W. Greene & P. G. M. Wuts, ProtectiveGroups in Organic Synthesis, John Wiley & Sons, 1991. The protectinggroups may be removed at a convenient subsequent stage usingconventional methods known from the art.

The method of treating neuronal disorders as described in the presentinvention, may also be carried out using a pharmaceutical compositioncomprising at least one effector of QC optionally in combination with atleast one PEP-inhibitor and/or at least one DP IV-inhibitor and/or atleast one NPY-receptor-ligand and/or at least one ACE-inhibitor or anyother of the compounds as defined herein and a pharmaceuticallyacceptable carrier. The pharmaceutical composition may contain betweenabout 0.01 mg and 100 mg, preferably about 5 to 50 mg, of each compound,and may be constituted into any form suitable for the mode ofadministration selected. Carriers include necessary and inertpharmaceutical excipients, including, but not limited to, binders,suspending agents, lubricants, flavorants, sweeteners, preservatives,dyes, and coatings. Compositions suitable for oral administrationinclude solid forms, such as pills, tablets, caplets, capsules (eachincluding immediate release, timed release and sustained releaseformulations), granules, and powders, and liquid forms, such assolutions, syrups, elixirs, emulsions, and suspensions. Forms useful forparenteral administration include sterile solutions, emulsions andsuspensions.

Advantageously, compounds of the present invention may be administeredin a single daily dose, or the total daily dosage may be administered individed doses of two, three or four times daily. Furthermore, compoundsfor the present invention can be administered in intranasal form viatopical use of suitable intranasal vehicles, or via transdermal skinpatches well known to those of ordinary skill in that art. To beadministered in the form of transdermal delivery system, the dosageadministration will, of course, be continuous rather than intermittentthroughout the dosage regimen.

For instance, for oral administration in the form of a tablet orcapsule, the active drug component can be combined with an oral,non-toxic pharmaceutically acceptable inert carrier such as ethanol,glycerol, water and the like. Moreover, when desired or necessary,suitable binders; lubricants, disintegrating agents and coloring agentscan also be incorporated into the mixture. Suitable binders include,without limitation, starch, gelatin, natural sugars such as glucose orbetalactose, corn sweeteners, natural and synthetic gums such as acacia,tragacanth or sodium oleate, sodium stearate, magnesium stearate, sodiumbenzoate, sodium acetate, sodium chloride and the like. Disintegratorsinclude, without limitation, starch, methyl cellulose, agar, bentonite,xanthan gum and the like.

The liquid forms in suitable flavored suspending or dispersing agentssuch as the synthetic and natural gums, for example, tragacanth, acacia,methyl-cellulose and the like. For parenteral administration, sterilesuspensions and solutions are desired. Isotonic preparations whichgenerally contain suitable preservatives are employed when intravenousadministration is desired.

The compounds or combinations of the present invention can also beadministered in the form of liposome delivery systems, such as smallunilamellar vesicles, large unilamellar vesicles, and multilamellarvesicles. Liposomes can be formed from a variety of phospholipids, suchas cholesterol, stearylamine or phosphatidylcholines.

Compounds or combinations of the present invention may also be deliveredby the use of monoclonal antibodies as individual carriers to which thecompound molecules are coupled. The compounds of the present inventionmay also be coupled with soluble polymers as targetable drug carriers.Such polymers can include polyvinylpyrrolidone, pyran copolymer,polyhydroxypropylmethacrylamidephenol,polyhydroxyethylaspartamid-ephenol, or polyethyleneoxidepolyllysinesubstituted with palmitoyl residue. Furthermore, the compounds of thepresent invention may be coupled to a class of biodegradable polymersuseful in achieving controlled release of a drug, for example, polyacticacid, polyepsilon caprolactone, polyhydroxy butyeric acid,polyorthoesters, polyacetals, polydihydropyrans, polycyanoacrylates andcross-linked or amphipathic block copolymers of hydrogels.

Compounds or combinations of this invention may be administered in anyof the foregoing compositions and according to dosage regimensestablished in the art whenever treatment of the addressed disorders isrequired.

The daily dosage of the products may be varied over a wide range from0.01 to 1.000 mg per mammal per day. For oral administration, thecompositions are preferably provided in the form of tablets containing,0.01, 0.05, 0.1, 0.5, 1.0, 2.5, 5.0, 10.0, 15.0, 25.0, 50.0, 100, 150,200, 250 and 500 milligrams of each active ingredient or combinationsthereof for the symptomatic adjustment of the dosage to the patient tobe treated. An effective amount of the drug is ordinarily supplied at adosage level of from about 0.1 mg/kg to about 300 mg/kg of body weightper day. Preferably, the range is from about 1 to about 50 mg/kg of bodyweight per day. The compounds or combinations may be administered on aregimen of 1 to 4 times per day.

Optimal dosages to be administered may be readily determined by thoseskilled in the art, and will vary with the particular compound used, themode of administration, the strength of the preparation, the mode ofadministration, and the advancement of disease condition. In addition,factors associated with the particular patient being treated, includingpatient age, weight, diet and time of administration, will result in theneed to adjust dosages.

Suitably, the particularly beneficial effect provided by the treatmentof the invention is an improved therapeutic ratio for the combination ofthe invention relative to the therapeutic ratio for one compound of thecombination when used alone and at a dose providing an equivalentefficacy to the combination of the invention.

In a preferred aspect, the particularly beneficial effect provided bythe treatment of the invention is indicated to be a synergistic effectrelative to the control expected from the effects of the individualactive agents.

In a further aspect of the invention, combining doses of at least oneQC-inhibitor with at least one PEP-inhibitor and/or at least one DPIV-inhibitor and/or at least one NPY-receptor-ligand will produce agreater beneficial effect than can be achieved for either agent alone ata dose twice that used for that agent in the combination.

In a preferred aspect, the dosage level of each of the active agentswhen used in accordance with the treatment of the invention will be lessthan would have been required from a purely additive effect upon theneuronal condition.

It is also considered that the treatment of the invention will effect animprovement, relative to the individual agents, in decreasing theintracellular deposition of pGlu-amyloid-β-peptides and therebydramatically slowing down the plaque formation in the brain of a mammal,preferably in human brain.

In a further aspect, the invention also provides a process for preparinga pharmaceutical composition comprising at least one at least oneeffector of QC optionally in combination with at least one PEP-inhibitorand/or at least one DP IV-inhibitor and/or at least oneNPY-receptor-ligand and/or at least one ACE-inhibitor and apharmaceutically acceptable carrier therefor, which process comprisesadmixing the QC effector and/or DP IV-inhibitor and/or the PEP-inhibitorand/or the NPY-receptor-ligand and/or the ACE-inhibitor and apharmaceutically acceptable carrier.

The compositions are preferably in a unit dosage form in an amountappropriate for the relevant daily dosage.

Suitable dosages, including especially unit dosages, of theQC-inhibitor, the PEP-inhibitor, the DP IV-inhibitor and theNPY-receptor-ligand include the known dosages including unit doses forthese compounds as described or referred to in reference text such asthe British and US Pharmacopoeias, Remington's Pharmaceutical Sciences(Mack Publishing Co.), Martindale The Extra Pharmacopoeia (London, ThePharmaceutical Press) (for example see the 31st Edition page 341 andpages cited therein) or the above mentioned publications.

EXAMPLES OF THE INVENTION Example 1 Solid-Phase Synthesis of Peptides

The peptides used herein were synthesized with an automated synthesizerSYMPHONY (RAININ) using a modified Fmoc-protocol. Cycles were modifiedby using double couplings from the 15^(th) amino acid from theC-terminus of the peptide with five-fold excess of Fmoc-amino acids andcoupling reagent. The peptide couplings were performed byTBTU/NMM-activation using a 0.23 mmol substituted NovaSyn TGR-resin orthe corresponding preloaded Wang-resin at 25 μmol scale. The cleavagefrom the resin was carried out by a cleavage-cocktail consisting of94.5% TFA, 2.5% water, 2.5% EDT and 1% TIS.

Analytical and preparative HPLC were performed by using differentgradients on the LiChrograph HPLC system of Merck-Hitachi. The gradientswere made up from two solvents: (A) 0.1% TFA in H₂O and (B) 0.1% TFA inacetonitrile. Analytical HPLC were performed under the followingconditions: solvents were run (1 ml/min) through a 125-4 NucleosilRP18-column, over a gradient from 5%-50% B over 15 min and then up to95% B until 20 min, with UV detection (λ=220 nm). Purification of thepeptides was carried out by preparative HPLC on either a 250-20Nucleosil 100 RP8-column or a 250-10 LiChrospher 300 RP18-column (flowrate 6 ml/min, 220 nm) under various conditions depending on peptidechain length.

For the identification of the peptides and peptide analogues, laserdesorption mass spectrometry was employed using the HP G2025 MALDI-TOFsystem of Hewlett-Packard.

Example 2 Determination of IC₅₀-Values of DP IV-Inhibitors

100 μl inhibitor stock solution were mixed with 100 μl buffer (HEPES pH7.6) and 50 μl substrate (Gly-Pro-pNA, final concentration 0.4 mM) andpreincubated at 30° C. Reaction was started by addition of 20 μlpurified porcine DP IV. Formation of the product pNA was measured at 405nm over 10 min using the HTS 7000Plus plate reader (Perkin Elmer) andslopes were calculated. The final inhibitor concentrations rangedbetween 1 mM and 30 nM.

For calculation of IC₅₀-values GraFit 4.0.13 (Erithacus Software) wasused.

Example 3 Determination of K_(i)-values of DP IV-inhibitors

For determination of the K_(i)-values DP IV activity was measured in thesame way as described in example 2 at final substrate concentrations of0.05, 0.1, 0.2, and 0.4 mM and further 7 inhibitor concentrationscovering the IC₅₀ concentration. Calculations were performed using theGraFit Software.

Example 4 Prolyl endopeptidase (PEP) enzymatic activity assays

The enzymatic activity of PEP was quantified as described recently(Schulz et al., 2002, Modulation of inositol 1,4,5-triphosphateconcentration by prolyl endopeptidase inhibition. Eur J Biochem 269:5813-5820). Cellular extracts as described above were incubated in theassay buffer using the fluorogenic substrate Z-Gly-Pro-NHMec (10 μM;Bachem, Heidelberg, Germany) on a spectrofluorimeter SFM 25 (excitationwavelength 380 nm, emission wavelength 460 nm, Kontron, Neufahrn,Germany) equipped with a four-cell changer and controlled by anIBM-compatible personal computer. The data obtained were analyzed withthe software FLUCOL (Machleidt et al., 1995).

Example 5 Assays for Glutaminyl Cyclase Activity Fluorometric Assays

All measurements were performed with a BioAssay Reader HTS-7000Plus formicroplates (Perkin Elmer) at 30° C. QC activity was evaluatedfluorometrically using H-Gln-βNA. The samples consisted of 0.2 mMfluorogenic substrate, 0.25 U pyroglutamyl aminopeptidase (Unizyme,Hørsholm, Denmark) in 0.2 M Tris/HCl, pH 8.0 containing 20 mM EDTA andan appropriately diluted aliquot of QC in a final volume of 250 μl.Excitation/emission wavelengths were 320/410 nm. The assay reactionswere initiated by addition of glutaminyl cyclase. QC activity wasdetermined from a standard curve of β-naphthylamine under assayconditions. One unit is defined as the amount of QC catalyzing theformation of 1 μmol pGlu-βNA from H-Gln-βNA per minute under thedescribed conditions.

In a second fluorometric assay, QC was activity was determined usingH-Gln-AMC as substrate. Reactions were carried out at 30° C. utilizingthe NOVOStar reader for microplates (BMG labtechnologies). The samplesconsisted of varying concentrations of the fluorogenic substrate, 0.1 Upyroglutamyl aminopeptidase (Qiagen) in 0.05 M Tris/HCl, pH 8.0containing 5 mM EDTA and an appropriately diluted aliquot of QC in afinal volume of 250 μl. Excitation/emission wavelengths were 380/460 nm.The assay reactions were initiated by addition of glutaminyl cyclase. QCactivity was determined from a standard curve of7-amino-4-methylcoumarin under assay conditions. The kinetic data wereevaluated using GraFit software.

Spectrophotometric Assay of QC

This novel assay was used to determine the kinetic parameters for mostof the QC substrates. QC activity was analyzed spectrophotometricallyusing a continuous method, that was derived by adapting a previousdiscontinuous assay (Bateman, R. C. J. 1989 J Neurosci Methods 30,23-28) utilizing glutamate dehydrogenase as auxiliary enzyme. Samplesconsisted of the respective QC substrate, 0.3 mM NADH, 14 mMα-Ketoglutaric acid and 30 U/ml glutamate dehydrogenase in a finalvolume of 250 μl. Reactions were started by addition of QC and persuedby monitoring of the decrease in absorbance at 340 nm for 8-15 min.

The initial velocities were evaluated and the enzymatic activity wasdetermined from a standard curve of ammonia under assay conditions. Allsamples were measured at 30° C., using either the SPECTRAFluor Plus orthe Sunrise (both from TECAN) reader for microplates. Kinetic data wasevaluated using GraFit software.

Inhibitor Assay

For inhibitor testing, the sample composition was the same as describedabove, except of the putative inhibitory compound added. For a rapidtest of QC-inhibition, samples contained 4 mM of the respectiveinhibitor and a substrate concentration at 1 K_(M). For detailedinvestigations of the inhibition and determination of K_(i)-values,influence of the inhibitor on the auxiliary enzymes was investigatedfirst. In every case, there was no influence on either enzyme detected,thus enabling the reliable determination of the QC inhibition. Theinhibitory constant was evaluated by fitting the set of progress curvesto the general equation for competitive inhibition using GraFitsoftware.

Example 6 MALDI-TOF Mass Spectrometry

Matrix-assisted laser desorption/ionization mass spectrometry wascarried out using the Hewlett-Packard G2025 LD-TOF System with a lineartime of flight analyzer. The instrument was equipped with a 337 nmnitrogen laser, a potential acceleration source (5 kV) and a 1.0 mflight tube. Detector operation was in the positive-ion mode and signalswere recorded and filtered using LeCroy 9350M digital storageoscilloscope linked to a personal computer. Samples (5 μl) were mixedwith equal volumes of the matrix solution. For matrix solution we usedDHAP/DAHC, prepared by solving 30 mg 2′,6′-dihydroxyacetophenone(Aldrich) and 44 mg diammonium hydrogen citrate (Fluka) in 1 mlacetonitrile/0.1% TFA in water (1/1, v/v). A small volume (≈1 μl) of thematrix-analyte-mixture was transferred to a probe tip and immediatelyevaporated in a vacuum chamber (Hewlett-Packard G2024A sample prepaccessory) to ensure rapid and homogeneous sample crystallization. Forlong-term testing of Glu¹-cyclization, Aβ-derived peptides wereincubated in 100 μl 0.1 M sodium acetate buffer, pH 5.2 or 0.1 MBis-Tris buffer, pH 6.5 at 30° C. Peptides were applied in 0.5 mM[Aβ(3-11)a] or 0.15 mM [Aβ(3-21)a] concentrations, and 0.2 U QC wasadded all 24 hours. In case of Aβ(3-21)a, the assays contained 1% DMSO.At different times, samples were removed from the assay tube, peptidesextracted using ZipTips (Millipore) according to the manufacturer'srecommendations, mixed with matrix solution (1:1 v/v) and subsequentlythe mass spectra recorded. Negative controls did either contain no QC orheat deactivated enzyme. For the inhibitor studies the samplecomposition was the same as described above, with exception of theinhibitory compound added (5 mM benzimidazole or 2 mM1,10-phenanthroline).

The first QC inhibitors are disclosed in WO 200409859. There are noother potent QC inhibitors known in the art. The same holds true forcombinations and compositions for the treatment of neuronal diseasescomprising QC inhibitors. Compounds and combinations of the inventionmay have the advantage that they are, for example, more potent, moreselective, have fewer side-effects, have better formulation andstability properties, have better pharmacokinetic properties, be morebioavailable, be able to cross blood brain barrier and are moreeffective in the brain of mammals, are more compatible or effective incombination with other drugsor be more readily synthesized than othercompounds of the prior art.

Throughout the specification and the claims which follow, unless thecontext requires otherwise, the word ‘comprise’, and variations such as‘comprises’ and ‘comprising’, will be understood to imply the inclusionof a stated integer, step, group of integers or group of steps but notto the exclusion of any other integer, step, group of integers or groupof steps.

All patents and patent applications mentioned above are hereinincorporated in their entirety by reference.

The invention embraces all combinations of preferred and more preferredgroups and embodiments of groups recited above.

1. A compound of the formula 1 including all stereoisomers orpharmaceutically acceptable salts thereof:

wherein: A is either: an alkyl chain, an alkenyl chain, an alkynyl chainor is a group selected from formulae (I) to (V):

wherein: R⁶, R⁷, R⁸, R⁹ and R¹⁰ are independently H or an alkyl chain,alkenyl chain, alkynyl chain, cycloalkyl, a carbocycle, aryl,heteroaryl, or a heterocycle; n and n¹ are independently 1-5; m is 1-5;o is 0-4; B is a group selected from formulae (VI) to (VIb):

wherein: D represents an alkyl chain, alkenyl chain, alkynyl chain, acycloalkyl, carbocycle, aryl, -alkylaryl, heteroaryl, -alkylheteroaryl,acyl or a heterocycle. X represents O or NR¹⁹, with the proviso forformulas (VIII) and (IX) that, if Z=CH, X is O or S; R¹⁹ is selectedfrom the group consisting of H, alkyl, cycloalkyl, aryl, heteroaryl,-oxyalkyl, -oxyaryl, carbonyl, amido, hydroxy, NO₂, NH₂, CN; with theproviso that the following compounds:

are excluded from formula
 1. 2. The compound according to claim 1,wherein A is an unbranched C₃ alkyl chain.
 3. The compound according toclaim 1, wherein A is a group of formula (I), (II) or (III), and n andn¹ are each
 1. 4. The compound according to any claim 1, wherein B isgroup (VI).
 5. The compound according to claims 1, wherein B is group(VIa).
 6. The compound according to claim 1, wherein X represents NR¹⁹.7. The compound according to claim 1, wherein X represents O.
 8. Thecompound according to claim 1, wherein D represents substituted phenyl.9. The compound according to claim 1, wherein D represents3,4-dimethoxyphenyl.
 10. The compound according to claim 1 correspondingto either formula 1b:

wherein R¹ and R² are selected from: R¹ R² Cyano Methyl Cyano3,4-(dimethoxy)-phenyl Cyano 2,4-(dimethoxy)-phenyl Cyano3,5-(dimethoxy)-phenyl Cyano 2,3-dihydrobenzo[b][1,4]dioxin-7-yl CyanoBenzo[d][1,3]dioxol-6-yl Cyano 3-(methoxy)-phenyl Cyano4-(ethoxy)-phenyl Cyano 4-(benzyloxy)-phenyl Cyano Phenyl Cyano4-(methoxy)-phenyl Cyano 4-(acetyl)-phenyl Cyano 4-(nitro)-phenyl CyanoBenzyl Cyano Naphthalen-1-yl Cyano 4-(fluoro)-phenyl Cyano4-(iodo)-phenyl Cyano 4-(bromo)-phenyl Cyano Cyclooctyl Cyano tert-butylCyano 4-(methyl)-phenyl Cyano 4-(methylthio)-phenyl Cyano4-(ethyl)-phenyl Cyano 4-(dimethylamino)-phenyl Cyano Butyl Cyano TritylCyano (Benzo[d][1,3]dioxol-6yl)methyl Cyano (tetrahydrofuran-2yl)methylCyano 4-(trifluoromethyl)-phenyl Cyano (furan-2-yl)methyl Cyano2-(morpholin-4-yl)-ethyl Cyano 2,3,4-(trimethoxy)-phenyl Cyano4-(oxazol-5yl)-phenyl Cyano Pyridin-3-yl Cyano 4-(cyano)-phenyl Cyano4-(trifluoromethoxy)-phenyl Cyano 4-(piperidinosulfonyl)-phenyl Cyano4-(1H-pyrazol-1-yl)phenyl H 3,4-(dimethoxy)-phenyl Methyl3,4-(dimethoxy)-phenyl Cyano Cycloheptyl Cyano 3,4,5-(trimethoxy)-phenyl

or formula 1c:

wherein R³ is selected from: R³ Ethyl 6-fluoro-4H-benzo[d][1,3]dioxin-8-yl 3-(cylopentyloxy)-4-(methoxy)-phenyl4-(heptyloxy)-phenyl 4-(butoxy)-phenyl 3,4-(dimethoxy)-phenyl3,4-dihydro-2H-benzo[b] [1,4]dioxepin-7-yl

or the following compound:

or a pharmaceutically acceptable salt or stereoisomers thereof.
 11. Apharmaceutical composition comprising at least one compound according toclaim 1 optionally in combination with a therapeutically acceptablecarrier and/or excipient.
 12. The pharmaceutical composition accordingto claim 11 for parenteral, enteral or oral administration.
 13. Apharmaceutical composition comprising at least one compound according toclaim 1 and at least one compound selected from the group consisting ofPEP-inhibitors, LiCl, inhibitors of dipeptidyl aminopeptidases,preferably inhibitors of DP IV or DP IV-like enzymes, NPY-receptorligands, NPY agonists, ACE inhibitors, PIMT enhancers, inhibitors ofbeta secretases, inhibitors of gamma secretases, inhibitors of neutralendopeptidase, PDE-4 inhibitors, MAO inhibitors, TNFalpha inhibitors,amyloid protein or amyloid peptide deposition inhibitors, sigma-1receptor inhibitors and histamine H3 antagonists.
 14. The pharmaceuticalcomposition according to claim 13, wherein said inhibitor of DP IV/DPIV-like enzymes is selected from the group consisting ofL-threo-isoleucyl pyrrolidide, L-allo-isoleucyl thiazolidide,L-allo-isoleucyl pyrrolidide and salts thereof, valine pyrrolidide,BMS-477118, CP-867534-01, LAF-237, PHX-1004, SSR-162369, SYR-322,TSL-225, FE-999011 GW-229A, 815541, K-579, MK-431, PT-100 and one of


15. The pharmaceutical composition according to claim 13, wherein saidNPY antagonist is selected from 3a,4,5,9b-tetrahydro-1h-benz[e]indol-2-yl amine, BIBP3226 and(R)—N2-(diphenylacetyl)-(R)—N-[1-(4-hydroxy-phenyl)ethyl] arginineamide.
 16. The pharmaceutical composition according to claim 13, whereinsaid PEP-inhibitor is selected from the group consisting of chemicalderivatives of proline or small peptides containing terminal prolines,benzyloxycarbonyl-prolyl-prolinal, N-terminal substituted L-proline,L-prolylpyrrolidine, substituted N-benzyloxycarbonyl (Z) dipeptidescontaining prolinal at the carboxy terminus, substituted thioprolines,substituted thiazolidines, substituted oxopyrrolidines, carboxy terminalmodified prolines including fluorinated ketone derivatives, chloromethylketone derivatives of acyl-proline or acylpeptide-proline(Z-Gly-Pro-CH₂Cl) and 2-acylpyrrolidine derivatives. or wherein saidPEP-inhibitor is selected from the group consisting of Fmoc-Ala-Pyrr-CN,Z-321, ONO-1603, JTP-4819, S-17092 and


17. The pharmaceutical composition according to claim 13, wherein saidACE-inhibitor is SDZ ENA 713 (rivastigmine(+)-(S)—N-ethyl-3-[(1-dimethylamino)ethyl]-N-methylphenylcarbamatehydrogen tartrate.
 18. The pharmaceutical composition according to claim13, wherein said PDE-4 inhibitor is selected from the group consistingof Rolipram, CC-002, L-826141, Sch-351591 (D-4396), OS-0217,IBFB-130011, IBFB-150007, IBFB-130020, IBFB-140301, IC-485, VMX-554,VMX-565, MEM-1414, MEM-1018, MEM-1091, MEM-1145, CI-1044, BHN, ZK-117137and SB-207499 or analogs thereof.
 19. The pharmaceutical compositionaccording to claim 13, wherein said PIMT enhancer is a10-aminoaliphatyl-dibenz[b,f]oxepine of the general formula

wherein alk is a divalent aliphatic radical, R is an amino group that isunsubstituted or mono- or di-substituted by monovalent aliphatic and/oraraliphatic radicals or disubstituted by divalent aliphatic radicals,and R₁, R₂, R₃ and R₄ are each, independently of the others, hydrogen,lower alkyl, lower alkoxy, halogen or trifluoromethyl.
 20. Thepharmaceutical composition according to claim 13, wherein said gammasecretase inhibitor is


21. The pharmaceutical composition according to claim 13, wherein saidbeta secretase inhibitor is


22. The pharmaceutical composition according to claim 13, wherein saidMAO inhibitor is ladostigil of the formula


23. The pharmaceutical composition according to claim 13, wherein saidhistamine H3 antagonist is a compound selected from the group consistingof A-331440, A-349821, 3874-H1, UCL-2173, UCL-1470, DWP-302,GSK-189254A, GSK-207040A, cipralisant, GT-2203,1S,2S)-2-(2-Aminoethyl)-1-(1H-imidazol-4-yl)cyclopropane, JNJ-5207852,NNC-0038-0000-1049, dual H1/H3, Sch-79687 and one of


24. A method of treatment of a neuronal disease selected fromAlzheimer's disease, Down Syndrome, Parkinson disease, ChoreaHuntington, pathogenic psychotic conditions, schizophrenia, impairedfood intake, sleep-wakefulness, impaired homeostatic regulation ofenergy metabolism, impaired autonomic function, impaired hormonalbalance, impaired regulation, body fluids, hypertension, fever, sleepdysregulation, anorexia, anxiety related disorders including depression,seizures including epilepsy, drug withdrawal or alcoholism,neurodegenerative disorders including cognitive dysfunction or dementia,which comprises administering to a mammal an effective amount of acompound of claim 1 including all stereoisomers or pharmaceuticallyacceptable salts thereof.
 25. The method according to claim 24 whereinthe neuronal disease is selected from Alzheimer's disease, DownSyndrome, Parkinson disease or Chorea Huntington.